Methods of purifying steviol glycosides and uses of the same

ABSTRACT

A method for purifying steviol glycosides by utilizing a plurality of columns is described.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent ApplicationNo. 61/577,202, filed Dec. 19, 2011, and U.S. Provisional PatentApplication No. 61/651,099, filed May 24, 2012, the contents of whichare incorporated by reference in their entirety.

FIELD OF THE INVENTION

The present invention relates generally to a process for purifying oneor more steviol glycosides, such as Rebaudioside X (Reb X), from asolution of steviol glycosides. The present invention also relates tosweetener compositions and sweetened compositions containing one or moresteviol glycosides, including Reb X, and methods for preparing the same.The present invention also relates to methods of providing a sugar-likeflavor and temporal profile to sweetener and sweetened compositionsutilizing Reb X.

BACKGROUND OF THE INVENTION

Natural caloric sugars, such as sucrose, fructose and glucose, areutilized to provide a pleasant taste to beverages, foods,pharmaceuticals, and oral hygienic/cosmetic products. Sucrose, inparticular, imparts a taste preferred by consumers. Although sucroseprovides superior sweetness characteristics, it is caloric. Non-caloricor low caloric sweeteners have been introduced to satisfy consumerdemand. However, sweeteners within this class differ from naturalcaloric sugars in ways that continue to frustrate consumers. On a tastebasis, non-caloric or low caloric sweeteners exhibit a temporal profile,maximal response, flavor profile, mouth feel, and/or adaptation behaviorthat differ from sugar. Specifically, non-caloric or low caloricsweeteners exhibit delayed sweetness onset, lingering sweet aftertaste,bitter taste, metallic taste, astringent taste, cooling taste and/orlicorice-like taste. On a source basis, many non-caloric or low caloricsweeteners are synthetic chemicals. The desire for a natural non-caloricor low caloric sweetener that tastes like sucrose remains high.

Stevia rebaudiana Bertoni is a perennial shrub of the Asteraceae(Compositae) family native to certain regions of South America. Itsleaves have been traditionally used for hundreds of years in Paraguayand Brazil to sweeten local teas and medicines. The plant iscommercially cultivated in Japan, Singapore, Taiwan, Malaysia, SouthKorea, China, Israel, India, Brazil, Australia and Paraguay.

The leaves of the plant contain a mixture containing diterpeneglycosides in an amount ranging from about 10 to 20% of the total dryweight. These diterpene glycosides are about 150 to 450 times sweeterthan sugar. Structurally, the diterpene glycosides are characterized bya single base, steviol, and differ by the presence of carbohydrateresidues at positions C13 and C19, as presented in FIGS. 2 a-2 k.Typically, on a dry weight basis, the four major steviol glycosidesfound in the leaves of Stevia are Dulcoside A (0.3%), Rebaudioside C(0.6-1.0%), Rebaudioside A (3.8%) and Stevioside (9.1%). Otherglycosides identified in Stevia extract include Rebaudioside B, D, E,and F, Steviolbioside and Rubusoside. Among these, only Stevioside andRebaudioside A are available on a commercial scale.

Steviol glycosides can be extracted from leaves using either water ororganic solvent extraction. Supercritical fluid extraction and steamdistillation methods have also been described. Methods for the recoveryof diterpene sweet glycosides from Stevia rebaudiana using supercriticalCO₂, membrane technology, and water or organic solvents, such asmethanol and ethanol, may also be used.

The use of steviol glycosides has been limited to date by certainundesirable taste properties, including licorice taste, bitterness,astringency, sweet aftertaste, bitter aftertaste, licorice aftertaste,and become more prominent with increase of concentration. Theseundesirable taste attributes are particularly prominent in carbonatedbeverages, where full replacement of sugar requires concentrations ofsteviol glycosides that exceed 500 mg/L. Use at that level results insignificant deterioration in the final product taste.

Accordingly, there remains a need to develop natural reduced ornon-caloric sweeteners that provide a temporal and flavor profilesimilar to that of sucrose.

There remains a further need to develop sweetened compositions, such asbeverages, that containing natural reduced or non-caloric sweetenersthat provide a temporal and flavor profile similar to that of sucrose.

SUMMARY OF THE INVENTION

The present invention provides a method for purifying the steviolglycoside Reb X from a solution of steviol glycosides:

In one embodiment, the present invention is a method for purifying Reb Xincluding passing a solution of steviol glycosides through amulti-column system including a plurality of columns packed with anadsorbent resin to provide at least one column having adsorbed steviolglycosides and eluting fractions with high Reb X content from the atleast one column having absorbed steviol glycosides to provide an elutedsolution with high Reb X content.

As the solution of steviol glycosides passes through the multi-columnsystem, the various steviol glycosides separate into different portionsof different columns. The portions differ from each other both by totalsteviol glycoside content and individual glycoside (particularly Reb X)content. Fractions containing high Reb X content are eluted/desorbedfrom the multi-column system separately from fractions containing lowReb X content.

Optionally, the method includes one or more additional steps. In oneembodiment, the method includes washing the multi-column system with awashing solution prior to eluting fractions with high Reb X content inorder to remove impurities.

In another embodiment, the method optionally includes decolorizing theeluted solution with high Reb X content, removing the alcohol solventand passing the remaining solution through a column with macroporousadsorbent to provide a second adsorption solution.

In another embodiment, the method optionally includes deionizing thesecond adsorption solution. The second adsorption solution can then beconcentrated to partially remove solvent to provide a high Reb X contentmixture containing from about 30% to about 40% solids content.

Further purification can be achieved by mixing a high Reb X contentmixture containing from about 30% to about 40% solids with a firstalcohol solvent to provide a Reb X solution, inducing crystallization toprovide first crystals of Reb X, and separating the first crystals ofReb X from the solution, wherein the first crystals have a purity levelgreater than about 60% (w/w) on a dry basis. In some embodiments, thepurity of the first crystals exceeds 60%, such as, for example, greaterthan about 65%, greater than about 70%, greater than about 75%, greaterthan about 80%, greater than about 85%.

To achieve higher purity levels, the first crystals may then besuspended in a second aqueous alcohol solution to provide secondcrystals of Reb X and a third aqueous alcohol solution. The secondcrystals of Reb X may be separated from the third aqueous alcoholsolution. These second crystals may have a purity level greater thanabout 90% (w/w) on a dry basis.

Fractions containing low Reb X content can also be further treatedaccording to certain methods provided herein. Optionally, the methodincludes one or more additional steps. In one embodiment, the methodincludes washing the multi-column system with a washing solution priorto eluting fractions with low Reb X content in order to removeimpurities.

In another embodiment, the method optionally includes decolorizing theeluted solution of steviol glycosides, removing the alcohol solvent andpassing the remaining solution through a column with macroporousadsorbent to provide a second adsorption solution.

In another embodiment, the method optionally includes deionizing theeluted solution of steviol glycosides. Removal of the remaining solventfrom the eluted solution—optionally decolorized and/ordeionized—provides a highly purified steviol glycoside mixture with atleast about 95% by weight total steviol glycosides on a dry basis.

The method of the present invention also includes preparing the solutionof steviol glycosides. In one embodiment, the solution of steviolglycosides is prepared by providing leaves of the Stevia rebaudianaBertoni plant, producing a crude extract by contacting the leaves withsolvent, separating insoluble material from the crude extract to providea first filtrate containing steviol glycosides, and treating the firstfiltrate to remove high molecular weight compounds and insolubleparticles, thereby providing a second filtrate containing steviolglycosides. The second filtrate is then treated with an ion-exchangeresin to remove salts, thereby providing a resin-treated filtrate thatserves as the solution of steviol glycosides in the method of thepresent invention.

The source of the solution of steviol glycosides may vary. In oneembodiment, the solution of steviol glycosides may be a commerciallyavailable stevia extract or steviol glycoside mixture. In anotherembodiment, the solution of steviol glycosides may be prepared fromplant material (e.g. leaves) of the Stevia rebaudiana Bertoni plant asdescribed herein. Alternatively, the solution of steviol glycosides maybe the by-product of another the isolation and purification processessteviol glycosides from Stevia rebaudiana Bertoni plant material.

According to one aspect of the invention, a method for producingpurified Reb X comprises the steps of: providing Stevia rebaudianaBertoni plant material; producing a crude extract by contacting theStevia rebaudiana Bertoni plant material with an extracting solvent,such as water; separating insoluble material from the first extract toprovide a filtrate containing steviol glycosides; deionizing thefiltrate; passing the filtrate feed over a series of columns packed withpolar macroporous resin and eluting steviol glycosides to provideeluates containing high Reb X and low Reb X fractions; decolorizing thesolutions; evaporating and deionizing; concentrating by nano-filters anddrying.

Sweetener compositions comprising Reb X are also provided herein. In oneembodiment, Reb X is present in an effective amount to provide asweetness equivalence from about 0.5 to about 14 degrees Brix of sucrosewhen present in a sweetened composition. In another embodiment, Reb X ispresent in an effective amount to provide a sucrose equivalence ofgreater than about 10% when present in a sweetened composition.

Reb X can be used in any form. In one embodiment, Reb X is the solesweetener in a sweetener composition. In another embodiment Reb X isprovided as part of a composition or mixture. In one embodiment, Reb Xis provided in a Stevia extract, wherein the Reb X component constitutesfrom about 5% to about 99% of the Stevia extract by weight on a drybasis. In a further embodiment, Reb X is provided in a mixture ofsteviol glycosides, wherein Reb X constitutes from about 5% to about 99%of the steviol glycoside mixture by weight on a dry basis.

The sweetener compositions can also contain one or more additionalsweeteners, including, for example, natural sweeteners, high potencysweeteners, carbohydrate sweeteners, synthetic sweeteners andcombinations thereof.

Particularly desirable sweetener compositions comprise Reb X and acompound selected from the group consisting of Reb A, Reb B, Reb D,NSF-02, mogroside V, erythritol or combinations thereof.

The sweetener compositions can also contain on or more additivesincluding, for example, carbohydrates, polyols, amino acids and theircorresponding salts, poly-amino acids and their corresponding salts,sugar acids and their corresponding salts, nucleotides, organic acids,inorganic acids, organic salts including organic acid salts and organicbase salts, inorganic salts, bitter compounds, flavorants and flavoringingredients, astringent compounds, proteins or protein hydrolysates,surfactants, emulsifiers, flavonoids, alcohols, polymers andcombinations thereof.

The sweetener compositions can also contain one or more functionalingredients, such as, for example, saponins, antioxidants, dietary fibersources, fatty acids, vitamins, glucosamine, minerals, preservatives,hydration agents, probiotics, prebiotics, weight management agents,osteoporosis management agents, phytoestrogens, long chain primaryaliphatic saturated alcohols, phytosterols and combinations thereof.

Methods of preparing sweetener compositions are also provided. In oneembodiment, a method for preparing a sweetener composition comprisescombining Reb X and at least one sweetener and/or additive and/orfunctional ingredient. In another embodiment, a method for preparing asweetener composition comprises combining a composition comprising Reb Xand at least one sweetener and/or additive and/or functional ingredient.

Sweetened composition containing Reb X or the sweetener compositions ofthe present invention are also provided herein. Sweetened compositionsinclude, for example, pharmaceutical compositions, edible gel mixes andcompositions, dental compositions, foodstuffs, beverages and beverageproducts.

Methods of preparing sweetened compositions are also provided herein. Inone embodiment, a method for preparing a sweetened composition comprisescombining a sweetenable composition and Reb X. The method can furtherinclude adding one or more sweetener, additive and/or functionalingredient. In another embodiment, a method for preparing a sweetenedcomposition comprises combining a sweetenable composition and asweetener composition comprising Reb X. The sweetener composition canoptionally comprise one or more sweetener, additive and/or functionalingredient.

In particular embodiments, beverages containing Reb X or the sweetenercompositions of the present invention are also provided herein. Thebeverages contain a liquid matrix, such as, for example, deionizedwater, distilled water, reverse osmosis water, carbon-treated water,purified water, demineralized water, phosphoric acid, phosphate buffer,citric acid, citrate buffer and carbon-treated water.

Full-calorie, mid-calorie, low-calorie and zero-calorie beveragescontaining Reb X or the sweetener compositions of the present inventionare also provided.

Methods of preparing beverages are also provided herein. In oneembodiment, a method for preparing a beverage comprises combining Reb Xand a liquid matrix. The method can further comprise adding one or moresweeteners, additives and/or functional ingredients to the beverage. Inanother embodiment, a method for preparing a beverage comprisescombining a sweetener composition comprising Reb X and a liquid matrix.

Tabletop sweetener compositions containing Reb X or the sweetenercompositions of the present invention are also provided herein. Thetabletop composition can further include at least one bulking agent,additive, anti-caking agent, functional ingredient and combinationsthereof. The tabletop sweetener composition can be present in the formof a solid or a liquid. The liquid tabletop sweetener can comprisewater, and optionally additives, such, as for example polyols (e.g.erythritol, sorbitol, propylene glycol or glycerol), acids (e.g. citricacid), antimicrobial agents (e.g. benzoic acid or a salt thereof).

Delivery systems comprising Reb X or the sweetener compositions of thepresent invention are also provided herein, such as, for example,co-crystallized sweetener compositions with a sugar or a polyol,agglomerated sweetener compositions, compacted sweetener compositions,dried sweetener compositions, particle sweetener compositions,spheronized sweetener compositions, granular sweetener compositions, andliquid sweetener compositions.

Finally, a method for imparting a more sugar-like temporal profile,flavor profile, or both to a sweetened composition comprises combining asweetenable composition with Reb X or the sweetener compositions of thepresent invention is also provided herein. The method can furtherinclude the addition of other sweeteners, additives, functionalingredients and combinations thereof.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings are included to provide a furtherunderstanding of the invention. The drawings illustrate embodiments ofthe invention and together with the description serve to explain theprinciples of the embodiments of the invention.

FIG. 1 shows the chemical structure of steviol glycosides in the Steviarebaudiana Bertoni leaves.

FIGS. 2 a-2 k show the chemical structures of Stevia rebaudiana Bertoniglycosides.

FIGS. 3 a, 3 b show HPLC traces of Reb X at various stages ofpurification. 3 a shows the HPLC trace of 80% pure Reb X. 3 b shows theHPLC trace of 97% Reb X (HPLC conditions provided in “Eluting theadsorbed steviol glycosides” section).

FIG. 4 shows the HPLC trace of Reb A, Reb B, Reb C, Reb D, Reb F,Stevioside, Dulcoside A, Steviolbioside and Rubusoside referencestandards (HPLC conditions provided in “Eluting the adsorbed steviolglycosides” section).

FIG. 5. shows the FTIR spectrum of Reb X.

FIG. 6 a, 6 b shows the high resolution spectral data for Reb X.

FIG. 7 a, 7 b. shows the ¹³C NMR spectrum of Reb X (150 MHz, C₅D₅N).

FIG. 8 a, 8 b, 8 c. shows the ¹H NMR spectrum of Reb X (600 MHz, C₅D₅N).

FIG. 9. shows the ¹H-¹H COSY spectrum of Reb X (600 MHz, C₅D₅N).

FIG. 10. shows the HMBC spectrum of Reb X (600 MHz, C₅D₅N).

FIG. 11. shows a sensory comparison of Reb X and Reb A in filteredwater.

FIG. 12. shows a sensory comparison of Reb X and Reb A in acidifiedwater.

FIG. 13. shows a sensory comparison of Reb X and NSF-02 at variousconcentrations in acidified water.

FIG. 14. shows a sensory comparison of Rob X and Reb B at variousconcentrations in acidified water.

FIG. 15. shows as sensory comparison of Reb X and Mogroside V at variousconcentrations in acidified water.

FIG. 16. shows a sensory comparison of Reb X and erythritol at variousconcentrations in acidified water.

FIG. 17. shows a sensory comparison of (i) Reb X, (ii) Rob X and Reb Aand (iii) Reb X and Reb D at various concentrations in acidified water.

FIG. 18. shows a sensory comparison of (i) Rob X, (ii) Reb X, Reb X andRob D and (iii) Reb X, Reb B and Reb D at various concentrations inacidified water.

FIG. 19. shows the chemical structure of Reb X.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the term “steviol glycoside(s)” refers to glycosides ofsteviol, including, but not limited to, naturally occurring steviolglycosides, e.g. Rebaudioside A, Rebaudioside B, Rebaudioside C,Rebaudioside D, Rebaudioside E, Rebaudioside F, Rebaudioside X,Stevioside, Steviolbioside, Dulcoside A, Rubusoside, etc. or syntheticsteviol glycosides, e.g. enzymatically glucosylated steviol glycosidesand combinations thereof.

As used herein, the term “total steviol glycosides” (TSG) is calculatedas the sum of the content of all steviol glycosides on a dry (anydrous)basis, including, for example, Rebaudioside A (Reb A), Rebaudioside B(Reb B), Rebaudioside C (Reb C), Rebaudioside D (Reb D), Rebaudioside E(Reb E), Rebaudioside F (Reb F), Rebaudioside X (Reb X), Stevioside,Steviolbioside, Dulcoside A and Rubusoside.

As used herein, the term “Reb X/TSG ratio” is calculated as the ratio ofReb X and TSG content on a dry basis as per the formula below:

{Reb X content(% dry basis)/TSG content(% dry basis)}×100%

As used herein, the term “solution of steviol glycosides” refers to anysolution containing a solvent and steviol glycosides. One example of asolution of steviol glycosides is the resin-treated filtrate obtainedfrom purification of Stevia rebaudiana plant material (e.g. leaves),described below, or by-products of other steviol glycosides' isolationand purification processes. Another example of a solution of steviolglycosides is a commercially available stevia extract brought intosolution with a solvent. Yet another example of a solution of steviolglycosides is a commercially available mixture of steviol glycosidesbrought into solution with a solvent.

In another aspect of the invention, a method for purifying Reb Xcomprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides; and    -   (b) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content.

In another aspect of the invention, a method for purifying Reb Xcomprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system; and    -   (c) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content;    -   (c) decolorizing the eluted solution with high Reb X content to        provide a first adsorption solution; and    -   (d) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content;    -   (d) decolorizing the eluted solution with high Reb X content to        provide a first adsorption solution; and    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin, to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content; and    -   (c) deionizing the solution.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content; and    -   (c) deionizing the solution.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content;    -   (c) decolorizing the eluted solution with high Reb X content to        provide a first adsorption solution;    -   (d) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution; and    -   (e) deionizing the second adsorption solution.

In another embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content;    -   (d) decolorizing the eluted solution with high Reb X content to        provide a first adsorption solution;    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution; and    -   (f) deionizing the second adsorption solution.

Removal of the alcoholic solvent from any of the above-mentionedprocesses relating to Reb X purification provides a high Reb X contentmixture. Subsequent removal of aqueous solvent provides a high Reb Xcontent mixture containing from about 30% to about 40% solids content,as discussed in the “Concentration” section below. Alternatively,substantially all of the solvent can removed to provide a dry powderwith high Reb X content.

In one embodiment, a method for purifying Reb X comprises:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with high Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution with high Reb X content;    -   (d) decolorizing the eluted solution with high Reb X content to        provide a first adsorption solution;    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution;    -   (f) deionizing the second adsorption solution; and    -   (g) removing the alcoholic solvent to provide a high Reb X        content mixture.

Further removal of aqueous solvents provides a high Reb X contentmixture containing from about 30% to about 40% solids content, asdiscussed in the “Concentration” section. Alternatively, substantiallyall of the solvent can removed to provide a dry powder with high Reb Xcontent.

In one embodiment, a method for purifying steviol glycosides includes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides; and    -   (b) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides.

In a more specific embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system; and    -   (c) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides.

In another embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides;    -   (c) decolorizing the eluted solution to provide a first        adsorption solution; and    -   (d) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution.

In a more specific embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides;    -   (d) decolorizing the eluted solution to provide a first        adsorption solution; and    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution.

In still another embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides; and    -   (c) deionizing the solution.

In a more specific embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractinos with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides; and    -   (d) deionizing the solution.

In yet another embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides;    -   (c) decolorizing the eluted solution to provide a first        adsorption solution;    -   (d) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution; and    -   (e) deionizing the second adsorption solution.

In a more specific embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides;    -   (d) decolorizing the eluted solution to provide a first        adsorption solution;    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution; and    -   (f) deionizing the second adsorption solution.

The eluted solution of steviol glycosides (decolorized and/or deionized)can be partially or fully dried, i.e. the solvent can be partially orcompletely removed to provide a semi- or entirely dry powder, asprovided below in the “Concentration” section. In one embodiment,complete removal of the solvent provides a purified mixture of steviolglycosides with total steviol glycoside content greater than about 95%on a dry basis.

In yet another embodiment, a method for purifying steviol glycosidesincludes:

-   -   (a) passing a solution of steviol glycosides through a        multi-column system including a plurality of columns packed with        an adsorbent resin to provide at least one column having        adsorbed steviol glycosides;    -   (b) removing impurities from the multi-column system;    -   (c) eluting fractions with low Reb X content from the at least        one column having adsorbed steviol glycosides to provide an        eluted solution of steviol glycosides;    -   (d) decolorizing the eluted solution to provide a first        adsorption solution;    -   (e) removing the alcoholic solvent from the first adsorption        solution and passing the remaining solution through a column        with a macroporous adsorbent to provide a second adsorption        solution;    -   (f) deionizing the second adsorption solution; and    -   (g) removing the solvent from the solution to provide a purified        steviol glycoside mixture with at least about 95% by weight        total steviol glycosides.

Preparing the Solution of Steviol Glycosides

Although the process for obtaining Reb X from Stevia rebaudiana leavesis provided herein, those of skill in the art will recognize that thetechniques described hereafter also apply to other starting materialscontaining Reb X, including, but not limited to, commercially availablestevia extracts, commercially available steviol glycoside mixtures,by-products of other steviol glycosides' isolation and purificationprocesses of the same.

Those of skill in the art will also recognize that certain stepsdescribed below, such as “separating insoluble material”, “removal ofhigh molecular weight compounds and insoluble particles” and “removingsalts” may be omitted when the starting materials do not containinsoluble material and/or high molecular weight compounds and/or salts.For example, in cases when already purified starting materials are used,such as commercially available stevia extracts, commercially availablesteviol glycoside mixtures, by-products of other steviol glycosides'isolation and purification processes of the same, one or more of theaforementioned steps may be omitted.

Those experienced in art will also understand that although the processdescribed below assumes certain order of the described steps this ordercan be altered in some cases.

The process of the present invention provides for complete retreatmentof Stevia rebaudiana Bertoni plant extract, with isolation andpurification of a highly purified steviol glycoside mixture or highlypurified individual sweet glycosides, such as Rebaudioside X. The plantextract can be obtained using any method such as, but not limited to,the extraction methods described in U.S. Pat. No. 7,862,845, the entirecontents of which are incorporated by reference herein, as well asmembrane filtration, supercritical fluid extraction, enzyme-assistedextraction, microorganism-assisted extraction, ultrasound-assistedextraction, microwave-assisted extraction, etc.

The solution of steviol glycosides may be prepared from Steviarebaudiana Bertoni leaves by contacting the Stevia rebaudiana Bertoniplant material with solvent to produce a crude extract, separatinginsoluble material from the crude extract to provide a first filtratecontaining steviol glycosides, treating the first filtrate to removehigh molecular weight compounds and insoluble particles, therebyproviding a second filtrate containing steviol glycosides and treatingthe second filtrate with an ion-exchange resin to remove salts toprovide a resin-treated filtrate.

In one embodiment, the Stevia rebaudiana plant material (e.g. leaves)may be dried at temperatures between about 20° C. to about 60° C. untila moisture content between about 5% and about 8% is reached. In aparticular embodiment, the plant material may be dried between about 20°C. and about 60° C. for a period of time from about 1 to about 24 hours,such as, for example, between about 1 to about 12 hours, between about 1to about 8 hours, between about 1 to about 5 hours or between about 2hours to about 3 hours. In other particular embodiments, the plantmaterial may be dried at temperatures between about 40° C. to about 45°C. to prevent decomposition.

In some embodiments, the dried plant material is optionally milled.Particle sizes may be between about 10 to about 20 mm.

The amount of Reb X in the plant material of the Stevia rebaudianaBertoni can vary. Generally speaking, Reb X should be present in anamount of at least about 0.001% by weight on an anhydrous basis.

The plant material (milled or unmilled) may be extracted by any suitableextraction process, such as, for example, continuous or batch refluxextraction, supercritical fluid extraction, enzyme-assisted extraction,microorganism-assisted extraction, ultrasound-assisted extraction,microwave-assisted extraction, etc. The solvent used for the extractioncan be any suitable solvent, such as for example, polar organic solvents(degassed, vacuumed, pressurized or distilled), non-polar organicsolvents, water (degassed, vacuumed, pressurized, deionized, distilled,carbon-treated or reverse osmosis) or a mixture thereof. In a particularembodiment, the solvent comprises water and one or more alcohols. Inanother embodiment, the solvent is water. In another embodiment, thesolvent is one or more alcohols.

In a particular embodiment, the plant material is extracted with waterin a continuous reflux extractor. One of skill in the art will recognizethe ratio of extraction solvent to plant material will vary based on theidentity of the solvent and the amount of plant material to beextracted. Generally, the ratio of extraction solvent to kilogram of dryplant material is from about 20 liters to about 25 liters to about onekilogram of leaves.

The pH of the extraction solvent can be between about pH 2.0 and 7.0,such as, for example, between about pH 2.0 and about pH 5.0, betweenabout pH 2.0 and about pH 4.0 or between about pH 2.0 and about pH 3.0.In a particular embodiment, the extraction solvent is aqueous, e.g.water and, optionally, acid and/or base in an amount to provide a pHbetween about pH 2.0 and 7.0, such as, for example, between about pH 2.0and about pH 5.0, between about pH 2.0 and about pH 4.0 or between aboutpH 2.0 and about pH 3.0. Any suitable acid or base can be used toprovide the desired pH for the extraction solvent, such as, for example,HCl, NaOH, citric acid, and the like.

The extraction may be carried out at temperatures between about 25° C.and about 90° C., such as, for example, between about 30° C. and about80° C., between about 35° C. and about 75° C., between about 40° C. andabout 70° C., between about 45° C. and about 65° C. or between about 50°C. and about 60° C.

In embodiments where the extraction process is a batch extractionprocess, the duration of extraction may range from about 0.5 hours toabout 24 hours, such as, for example, from about 1 hour to about 12hours, from about 1 hour to about 8 hours, or from about 1 hour to about6 hours.

In embodiments where the extraction process is a continuous process, theduration of extraction may range from about 1 hour to about 5 hours,such as, for example, from about 2.5 hours to about 3 hours.

After extraction, the insoluble plant material may be separated from thesolution by filtration to provide a filtrate containing steviolglycosides, referred to herein as a “first filtrate containing steviolglycosides”. Separation can be achieved by any suitable means including,but not limited to, gravity filtration, a plate-and-frame filter press,cross flow filters, screen filters, Nutsche filters, belt filters,ceramic filters, membrane filters, microfilters, nanofilters,ultrafilters or centrifugation. Optionally various filtration aids suchas diatomaceous earth, bentonite, zeolite etc, may also be used in thisprocess.

After separation, the pH of the first filtrate containing steviolglycosides may be adjusted to remove additional impurities. In oneembodiment, the pH of the first filtrate containing steviol glycosidescan be adjusted to between about 8.5 and about 10.0 by treatment with abase, such as, for example, calcium oxide or hydroxide (about 1.0% fromthe volume of filtrate) with slow agitation.

Treatment of the first filtrate with the base, as set forth above,results in a suspension, the pH of which can be adjusted to about 3.0 toabout 4.0 by treatment with any suitable flocculation/coagulation agent.Suitable flocculation/coagulation agents include, for example, potassiumalum, aluminum sulfate, aluminum hydroxide, aluminum oxide, CO₂, H₃PO₄,P₂O₅, MgO, SO₂, anionic polyacrylamides, quaternary ammonium compoundswith long-chain fatty acid substitutents, bentonite, diatomaceous earth,KemTab Sep series, Superfloc series, KemTab Flote series, Kemtalo Melseries, Midland PCS-3000, Magnafloc LT-26, Zuclar 100, Prastal 2935,Talofloc, Magox, soluble ferrous salts or a combination thereof.Exemplary ferrous salts include, but arc not limited to, FeSO₄, FeCl₂,Fe(NO₃)₃, Fe(SO₄)₃, FeCl₃ and combinations thereof. In a particularembodiment, the ferrous salt is FeCl₃. The filtrate may be treated withthe flocculation/coagulation agent for a duration of time between about5 minutes to about 1 hour, such as, for example, from about 5 minutes toabout 30 minutes, from about 10 minutes to about 20 minutes or fromabout 10 minutes to about 15 minutes. Stirred or slow agitation can alsobe used to facilitate treatment. Optionally, the pH of resultant mixturemay then be adjusted to between about 8.5 and about 9.0 with a base,such as, for example, calcium oxide or sodium hydroxide. The duration oftime for treatment with base, and optionally, with agitation, is betweenabout 5 minutes to about 1 hour, such as, for example, from about 10minutes to about 50 minutes, from about 15 minutes to about 45 minutes,from about 20 minutes to about 40 minutes or from about 25 minutes toabout 35 minutes. In a particular embodiment, the base is calcium oxideused for a between about 15 and about 40 minutes with slow agitation.

Precipitated high molecular weight compounds and insoluble particles areseparated from the mixture to provide second filtrate containing steviolglycosides. Separation can be achieved by any suitable means including,but not limited to, gravity filtration, a plate-and-frame filter press,cross flow filters, screen filters, Nutsche filters, belt filters,ceramic filters, membrane filters, microfilters, nanofilters,ultrafilters or centrifugation. Optionally various filtration aids suchas diatomaceous earth, bentonite, zeolite etc, may be used in thisprocess.

The second filtrate containing steviol glycosides may then be subjectedto preliminary deionization by any suitable method including, forexample, electrodialysis, filtration (nano- or ultra-filtration),reverse osmosis, ion exchange, mixed bed ion exchange or a combinationof such methods. In one embodiment, the second filtrate containingsteviol glycosides is deionized by treatment with one or more ionexchange resins to provide a resin-treated filtrate. In one embodiment,the second filtrate containing steviol glycosides is passed through astrong acid cation exchange resin. In another embodiment, the secondfiltrate containing steviol glycosides is passed through a weak baseanion-exchange resin. In still another embodiment, the second filtratecontaining steviol glycosides is passed through a strong acidcation-exchange resin followed by a weak base anion-exchange resin. Inyet another embodiment, the second filtrate containing steviolglycosides is passed through a weak base anion-exchange resin followedby a strong acid cation-exchange resin.

The cation-exchange resin can be any strong acid cation-exchanger wherethe functional group is, for example, sulfonic acid. Suitable strongacid cation-exchange resins are known in the art and include, but arenot limited to, Rohm & Haas Amberlite® 10 FPC22H resin, which is asulfonated divinyl benzene styrene copolymer, Dowex® ion exchange resinsavailable from Dow Chemical Company, 15 Serdolit® ion exchange resinsavailable from Serva Electrophoresis GmbH, T42 strong acidic cationexchange resin and A23 strong base an ion exchange resin available fromQualichem, Inc., and Lewatit strong ion exchange resins available fromLanxess. In a particular embodiment, the strong acid cation-exchangeresin is Amberlite® 10 FPC22H resin (H⁺). As would be known to thoseskilled in the art, other suitable strong acid cation-exchange resinsfor use with embodiments of this invention are commercially available.

The anion-exchange resin can be any weak base anion-exchanger where thefunctional group is, for example, a tertiary amine. Suitable weak baseanion exchange resins are known in the art and include, but are notlimited to, resins such as Amberlite-FPA53 (OH⁻), Amberlite IRA-67,Amberlite IRA-95, Dowex 67, Dowex 77 and Diaion WA 30 may be used. In aparticular embodiment, the strong acid cation-exchange resin isAmberlite-FPA53 (OH⁻) resin. As would be known to those skilled in theart, other suitable weak base anion-exchange resins for use withembodiments of this invention are commercially available.

In a particular embodiment, the second filtrate containing steviolglycosides is passed through a strong acid cation-exchange resin, e.g.Amberlite® 10 FPC22H resin (H⁺), followed by a weak base anion-exchangeresin, e.g. Amberlite-FPA53 (OH⁻), to provide a resin-treated filtrate.The specific velocity (SV) through one or more of the ion exchangecolumns can be between about 0.01 to about 5 hour⁻¹, such as, forexample between about 0.05 to about 4 hour⁻¹, between about 1 and about3 hour⁻¹ or between about 2 and about 3 hour⁻¹. In a particularembodiment, the specific velocity through the one or more ion exchangecolumns is about 0.8 hour⁻¹. Following completion of passing the secondfiltrate containing steviol glycosides through one or more ion exchangecolumns, the one or more ion exchange columns are washed with water,preferably reverse osmosis (RO) water. The solution obtained from thewater wash and the resin-treated filtrate may be combined beforeproceeding to the multi-column step.

Adsorption of the Solution of Steviol Glycosides

In certain embodiment, the solution of steviol glycosides is theresin-treated filtrate obtained from purification of Stevia rebaudianaleaf, described above. In another embodiment, the solution of steviolglycosides is a commercially available stevia extract dissolved in asolvent. In yet another embodiment, the solution of steviol glycosidesis a commercially available extract where insoluble material and/or highmolecular weight compounds and/or salts have been removed.

Reb X content in the solution of steviol glycosides may vary dependingon the source of the solution of steviol glycosides. For example, inembodiments where the source of steviol glycosides is plant material,the concentration of Reb X can be between about 5 ppm to about 50,000ppm, such as, for example, from about 10,000 ppm to about 50,000 ppm. Ina particular embodiment, the concentration of Reb X in the solution ofsteviol glycosides, where the source of steviol glycosides is plantmaterial, is from about 5 ppm to about 50 ppm.

In embodiments where the source is non-plant material, the concentrationof Reb X in the solution of steviol glycosides can also vary. Inexemplary embodiments, the concentration of Reb X in the solution ofsteviol glycosides can be between about 5 ppm to about 50,000 ppm, suchas, for example, from about 5,000 ppm to about 10,000 ppm.

The Reb X/TSG ratio in the solution of steviol glycosides will also varydepending on the source of the steviol glycosides. In one embodiment,the Reb X/TSG in the solution of steviol glycosides is from about 0.5%to about 99%, such as, for example, from about 0.5% to about 10%, fromabout 0.5% to about 20%, from about 0.5% to about 30%, from about 0.5%to about 40%, from about 0.5% to about 50%, from about 0.5% to about60%, from about 0.5% to about 70%, from about 0.5% to about 80%, fromabout 0.5% to about 90%. In more particular embodiments, the Reb X/TSGin the solution of steviol glycosides is from about 0.5% to about 5%.

The solution of steviol glycosides may be passed through one or moreconsecutively connected columns (connected serially or in parallel)packed with polar macroporous polymeric adsorbent to provide at leastone column having adsorbed steviol glycosides. In some embodiments, thenumber of columns can be greater than 3, such as, for example, 5columns, 6 columns, 7 columns, 8 columns, 9 columns, 10 columns, 11column, 12 columns, 13 columns, 14 columns or 15 columns. In aparticular embodiment, the resin-treated filtrate is passed through 7columns.

In certain embodiments, the first column in the sequence can be a“catcher column”, which is used to adsorb certain impurities, such assterebins, that have higher adsorption rates and faster desorption ratesthan most steviol glycosides. In some embodiments, the “catcher column”size can be about one-third the size of the remaining columns. The ratioof internal diameter to column height or so-called “diameter: heightratio” of the columns shall be between about 1:1 to about 1:100, suchas, for example, about 1:2, about 1:6, about 1:10, about 1:13, about1:16, or about 1:20. In a particular embodiment, the diameter: heightratio of the column is about 1:3. In yet another embodiment, thediameter: height ratio is about 1:8. In still another embodiment, thediameter: height ratio is about 1:15.

The polar macroporous polymeric adsorbent may be any macroporouspolymeric adsorption resins capable of adsorbing steviol glycosides,such as, for example, the Amberlite® XAD series (Rohm and Haas), Diaion®HP series (Mitsubishi Chemical Corp), Sepabeads® SP series (MitsubishiChemical Corp), Cangzhou Yuanwei YWD series (Cangzhou Yuanwei ChemicalCo. Ltd., China), or the equivalent. The individual columns may bepacked with the same resin or with different resins. The columns may bepacked with sorbent up to from about 75% to about 100% of their totalvolume.

In embodiments wherein the multi-column system is connected in parallel,the inlet of each column may connect to a separate feed source while theoutlet of each column connects to a separate receiver. The ratio of thevolume of the first column to the volume of the second column ispreferably in the range of about 1:1 to 1:10. The ratio of the volume ofthe last column to the volume of the previous, or penultimate, column ispreferably in the range of about 3:1 to 1:10. The columns may bemaintained at a temperature in the range of about 5-80° C., andpreferably in the range of about 15-25° C.

The solvent that carries the steviol glycoside solution through thecolumn system can comprise alcohol, water, or a combination thereof (anaqueous alcoholic solvent). The water to alcohol ratio (vol/vol) in theaqueous alcoholic solvent may be in the range of about 99.9:0.1 to about60:40, such as, for example, about 99:1 to about 90:10. The specificvelocity (SV) can be from about 0.3⁻¹ to about 1.5⁻¹, such as, forexample, about 1.0 hour⁻¹.

The alcohol can be selected from, for example, methanol, ethanol,n-propanol, 2-propanol, 1-butanol, 2-butanol and mixtures thereof.

Impurities and different steviol glycosides are retained in differentsections of the column system. Impurities with higher affinities to thesorbent are retained in the first column, impurities with loweraffinities to the sorbent are retained in the last column, and differentsteviol glycosides are retained in different sections of the system atdifferent concentrations, depending on their affinities to the sorbent.Generally Reb X is retained in later columns. “Columns” is usedinterchangeably herein with “fractions”, both of which refer to columns,or sections of columns with the desirable content (e.g. Reb X). As aresult, the initial mixture of steviol glycosides separates intodifferent portions retained on different columns. The portions differfrom each other both by total steviol glycoside content and individualglycoside (particularly Reb X) content.

Removing Impurities from the Multi-Column System

Upon complete passage through the one or more columns, the resins canoptionally be washed with a washing solution to remove impurities fromthe one or more columns. Suitable washing solutions include an aqueousor alcoholic solution, where the aqueous solution can contain anysuitable acid or base to arrive at the desired pH. The water to alcoholratio (vol/vol) in the aqueous alcoholic solution is in the range ofabout 99.9:0.1 to about 60:40. Multiple washes of the columns with thesame, or different, wash solutions can be performed, followed bywash(es) with water until the pH of the effluent from the one or morecolumns is about neutral (i.e., has a pH from about 6.0 to about 7.0).In a particular embodiment, the resins of the one or more columns iswashed sequentially with one volume of water, two volumes of NaOH, onevolume of water, two volumes of HCl, and finally with two volumes ofwater until it reached a neutral pH. The elution of impurities iscarried out either from each column separately (parallel connection) orfrom two or more consecutively (serially) connected columns.

Eluting the Adsorbed Steviol Glycosides

Desorption can be carried out with an aqueous alcohol solution. Suitablealcohols include methanol, ethanol, n-propanol, 2-propanol, 1-butanol,2-butanol and mixtures thereof. In a particular embodiment, the aqueousalcoholic solution can contain between about 30% to about 70% alcoholcontent, such as, for example, between about 40% to about 60%, about 50%to about 60%, about 51%, about 52%, about 53%, about 54%, about 55%,about 56%, about 57%, about 58% or about 59%. In a particularembodiment, the aqueous alcoholic solution contains between about 50% toabout 52% ethanol. A SV between about 0.5 hour⁻¹ to about 3.0 hour¹,such as, for example, between about 1.0 hour⁻¹ and about 1.5 hour⁻¹ canbe used. Desorption of the first “catcher column”, which is optional,can be carried out separately from the non-“catcher column” columns.

In one embodiment, fractions with high Reb X content are eluted with anaqueous alcohol solution to provide an eluted solution with high Reb Xcontent. “High Reb X content”, as used herein, refers to any materialwhich has a higher Reb X/TSG ratio compared to the solution of steviolglycosides prior to passing through the multi-column system. In oneembodiment, the Reb X/TSG ratio is greater than about 1% higher than theReb X/TSG ratio of the solution of steviol glycosides. In anotherembodiment, the Reb X/TSG ratio is greater than about 2% higher, about3% higher, about 4% higher, about 5% higher, about 10% higher, about 15%higher, about 20% higher, about 25% higher, about 30% higher, about 35%higher, about 40% higher, about 45% higher, about 50% higher, about 55%higher, about 60% higher about 65% higher about 70% higher, about 75%higher, about 80% higher, about 85% higher, about 90% higher or about95% higher. Generally speaking, the later columns will contain “high RebX content” fractions.

In a particular embodiment, the remaining columns (excluding the“catcher column”) can also be eluted with an aqueous alcohol solutionand their eluates combined to provide an eluted solution of steviolglycosides with low Reb X content. “Low Reb X content”, as used herein,refers to any material which has a lower Reb X/TSG ratio compared to thesolution of steviol glycosides prior to passing through the multi-columnsystem. “Low Reb X content” also refers to any material which has zeroReb X content. Generally speaking, the initial columns will contain “lowReb X content”.

The Reb X/TSG ratio can be determined experimentally by HPLC or HPLC/MS.For example, chromatographic analysis can be performed on a HPLC/MSsystem comprising an Agilent 1200 series (USA) liquid chromatographequipped with binary pump, autosampler, thermostatted columncompartment, UV detector (210 nm), and Agilent 6110 quadrupole MSdetector interfaced with Chemstation data acquisition software. Thecolumn can be a “Phenomenex Prodigy 5u ODS3 250×4.6 mm; 5 μm (P/No.00G-4097-E0)” column maintained at 40° C. The mobile phase can be 30:70(vol/vol.) acetonitrile and water (containing 0.1% formic acid) and theflow rate through the column can be 0.5 mL/min. The steviol glycosidescan be identified by their retention times in such a method, which aregenerally around 2.5 minutes for Reb D, around 2.9 minutes for Reb X,5.5 minutes for Reb A, 5.8 minutes for Stevioside, 7.1 minutes for RebF, 7.8 minutes for Reb C, 8.5 minutes for Dulcoside A, 11.0 minutes forRubusoside, 15.4 minutes for Reb B and 16.4 minutes for Steviolbioside.One of skill in the art will appreciate that the retention times for thevarious steviol glycosides given above can vary with changes in solventand/or equipment.

Those of skill in the art will also recognize that one or more of the“decolorizing”, “second adsorption” and “deionization” steps, describedbelow, may be omitted, e.g. where generally higher purity startingmaterial solutions of steviol glycosides are used. Those experienced inart will also understand that although the process described belowassumes certain order of the described steps, this order can be alteredin some cases.

Decolorizing

Decolorization can be achieved with any known method, such as, forexample, treatment with activated carbon. The quantity of the activatedcarbon can be from about 0.1% (wt/vol) to about 0.8% (wt/vol). In aparticular embodiment, the quantity of activated carbon is from about0.25% (wt/vol) to about 0.30% (wt/vol). The suspension may becontinuously agitated. The temperature of the treatment can be betweenabout 20° C. and about 30° C., such as, for example, about 25° C. Thetreatment can be for any duration sufficient to decolorize the elutedsolution, such as, for example, between about 20 minutes and about 3hours, between 20 minutes and about 2 hours, between about 30 minutesand 1.5 hours or between about 1 hour and about 1.5 hours. Followingtreatment, separation of used carbon can be conducted by any knownseparation means, such as, for example, gravity or suction filtration,centrifugation or plate-and-frame press filter.

The eluted solution with high Reb X content can optionally bedecolorized separately from the eluted solution of steviol glycosideswith low Reb X content.

Second Adsorption

The decolorized solution (also referred to herein as “the firstadsorption solution”) can be distilled or evaporated with vacuum toremove alcoholic solvent and then passed through macroporous adsorbentsecond time to provide a second adsorption solution. The secondadsorption solution contains aqueous solvent.

Deionization

Generally any type of strong acid cation-exchanger and weakanion-exchangers can be used at this stage. In one embodiment, theeluted solution (e.g. the eluted solution with high Reb Xcontent—optionally decolorized or the eluted solution of steviolglycosides—optionally decolorized) can be passed through a strong acidcation exchange resin. In another embodiment, the eluted solution ispassed through a weak base anion-exchange resin. In still anotherembodiment, the eluted solution is passed through a strong acidcation-exchange resin followed by a weak base anion-exchange resin. Inyet another embodiment, the eluted solution is passed through a weakbase anion-exchange resin followed by a strong acid cation-exchangeresin. Suitable strong acid cation-exchange columns, weak baseanion-exchange columns and flow rates are provided above with respect toproduction of the resin-treated filtrate. In a particular embodiment,the eluted solution can be passed through columns packed withcation-exchange resin Amberlite FPC22H (H⁺) followed with anion-exchangeresin Amberlite FPA53 (OH⁻).

In one embodiment, the second adsorption solution can be passed througha strong acid cation exchange resin. In another embodiment, the secondadsorption solution is passed through a weak base anion-exchange resin.In still another embodiment, the second adsorption solution is passedthrough a strong acid cation-exchange resin followed by a weak baseanion-exchange resin. In yet another embodiment, the second adsorptionsolution is passed through a weak base anion-exchange resin followed bya strong acid cation-exchange resin. Suitable strong acidcation-exchange columns, weak base anion-exchange columns and flow ratesare provided above with respect to production of the resin-treatedfiltrate. In a particular embodiment, the second adsorption solution canbe passed through columns packed with cation-exchange resin AmberliteFPC22H (H⁺) followed with anion-exchange resin Amberlite FPA53 (OH⁻).

Those experienced in art will recognize that deionization may bealternatively conducted by means of mixed bed ion exchange,electrodialysis or various membranes such as, for example, reverseosmosis membranes, nanofiltration membranes or ultrafiltrationmembranes.

Concentration

The eluted solution (e.g. the eluted solution with high Reb Xcontent—optionally decolorized and/or deionized, the eluted solution ofsteviol glycosides—optionally decolorized and/or deionized) or thesecond adsorption solution (optionally deionized) can be distilled orevaporated with vacuum to remove alcoholic solvent.

Once the alcoholic solvent is removed, the remaining aqueous solventfrom the concentrate of steviol glycosides, or concentrated secondadsorption solution, can be removed by any suitable means, including,but not limited to, evaporation or vacuum, to provide a dry purifiedsteviol glycoside mixture with greater than 95% by weight total steviolglycosides on a dry basis.

Removal of alcoholic solvents from the eluted solution with high Reb Xcontent provides a high Reb X content mixture. Further concentration toremove aqueous solvent can then be carried out by any suitable method,such as, for example, nano-filtration or evaporation under reducedpressure conditions to provide a high Reb X content mixture containingfrom about 30% to about 40% solids content, such as, for example, fromabout 30% to about 35% solids content or from about 33% to about 35%solids content. The high Reb X content mixture containing from about 30%to about 40% solids content contains aqueous solvent.

Alternatively, all solvent from the eluted solution with high Reb Xcontent can be removed by any suitable method, such as, for example,nano-filtration or evaporation under reduced pressure, freeze drying,flash drying, spray drying or a combination thereof to provide a drypowder with high Reb X content.

Purification of Reb X

In one embodiment, purification of Reb X from a high Reb X contentmixture containing from about 30% to about 40% solids content can beachieved by mixing a high Reb X content mixture containing from about30% to about 40% solids content with a first alcoholic solvent toprovide a Reb X solution and inducing crystallization. Generally, theratio of solvent to solids is from about 0.5 liters to about 100 litersper one kilogram of solid. In particular embodiments, the ratio ofsolvent to solids can be from about 3 to about 10 liters of solvent perone kilogram of solid. The alcohol can be any suitable alcohol, such as,for example, methanol, ethanol, n-propanol, 2-propanol, 1-butanol,2-butanol and mixtures thereof. The alcohol can contain small amounts ofwater or be anhydrous. In a particular embodiment, the alcohol isanhydrous methanol.

In another embodiment, purification of a high Reb X content mixturecontaining more than about 40% solids content can be achieved bydiluting the mixture with water to provide a high Reb X content mixturecontaining from about 30% to about 40% solids content, mixing themixture with an alcoholic solvent to provide a Reb X solution andinducing crystallization.

In yet another embodiment, a dry powder with high Reb X content can bemixed with an aqueous alcoholic solvent to provide a Reb X solution(preferably containing from about 30% to about 40% solids content) andinducing crystallization.

To induce crystallization, the Reb X solution is maintained at atemperature between about 20° C. and about 25° C., such as, for example,between about 20° C. and about 22° C., and, if necessary, seeded withReb X crystals. The duration of mixing can be between about 1 hour andabout 48 hours, such as, for example, about 24 hours.

Reb X crystals having a purity greater than about 60% by weight on a drybasis (referred to herein as the “first crystals of Reb X”) in a mixtureof steviol glycosides can be obtained after separation of the crystalsfrom the solution. In a particular embodiment, Reb X with a puritygreater than about 60%, about 65%, about 75%, about 80%, about 85%,about 90% or about 95% is obtained by this process.

Those of skill in the art will recognize that the purity of the firstcrystals of Reb X will depend on the Reb X content of the initialsolution of steviol glycosides among other variables. Accordingly, ifneeded, further wash steps can be performed to provide Reb X crystalswith higher purity. To produce Reb X with greater purity, the firstcrystals of Reb X can combined with a aqueous alcohol solution (referredto herein as the “second aqueous alcohol solution”) to provide secondcrystals of Reb X and a third aqueous alcohol solution. Separation ofthe second crystals of Reb X crystals from the third aqueous alcoholsolution provides second crystals of Reb X having a purity greater thanabout 90% by weight on a dry basis. In certain embodiments, Reb X withpurities greater than about 91%, about 92%, about 93%, about 94%, about95%, about 96%, about 97%, about 98% or about 99% can be obtained. Thisprocess can be repeated, as necessary, until the desired purity level isachieved. The cycle can be repeated two times, three times, four timesor five times. In some embodiments, water can be used instead of anaqueous alcohol solution.

The solution or suspension can be maintained at temperatures betweenabout 40° C. to about 75° C., such as, for example, from about 50° C. toabout 60° C. or about 55° C. to about 60° C. The duration that themixture can be maintained at a temperature between about 40° C. to about75° C. may vary, but can last between about 5 minutes and about 1 hour,such as, for example, between about 15 and about 30 minutes. The mixturecan then be cooled to a temperature between about 20° C. to about 22°C., for example. The duration that the mixture can be maintained at thecool temperature may vary, but can last between about 1 hour and about 5hours, such as, for example, between about 1 hour and about 2 hours.Agitation can optionally be used during the wash cycle.

Separation of Reb X crystals from the solution or suspension can beachieved by any known separation method, including, but not limited to,centrifugation, gravity or vacuum filtration, or drying. Different typeof dryers such as fluid bed dryers, rotary tunnel dryers, or platedryers may be used.

In some embodiments, when Reb X crystals are combined with water oraqueous alcohol solution, the Reb X may dissolve and accumulate inliquid phase. In that case the higher purity Reb X crystals may beobtained by drying or evaporative crystallization of liquid phase.

Sweetener Compositions

Sweetener compositions, as used herein, mean compositions that containat least one sweet component in combination with at least one othersubstance, such as, for example, another sweetener or an additive.

Sweetenable compositions, as used herein, mean substances which arecontacted with the mouth of man or animal, including substances whichare taken into and subsequently ejected from the mouth and substanceswhich are drunk, eaten, swallowed or otherwise ingested, and are safefor human or animal consumption when used in a generally acceptablerange.

Sweetened compositions, as used herein, mean substances that containboth a sweetenable composition and a sweetener or sweetener composition.

For example, a beverage with no sweetener component is a type ofsweetenable composition. A sweetener composition comprising Reb X anderythritol can be added to the un-sweetened beverage, thereby providinga sweetened beverage. The sweetened beverage is a type of sweetenedcomposition.

The sweetener compositions of the present invention include Reb X(13-[2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl)oxy]entkaur-16-en-19-oic acid-[2-O-β-D-glucopyranosyl-3-O-β-D-glycopyranosyl)ester having the formula:

Reb X may be provided in a purified form or as a component of a mixturecontaining Reb X and one or more additional components (i.e. a sweetenercomposition comprising Reb X). In one embodiment, Reb X is provided as acomponent of a mixture. In a particular embodiment, the mixture is aStevia extract. The Stevia extract may contain Reb X in an amount thatranges from about 5% to about 99% by weight on a dry basis, such as, forexample, from about 10% to about 99%, from about 20% to about 99%, fromabout 30% to about 99%, from about 40% to about 99%, from about 50% toabout 99%, from about 60% to about 99%, from about 70% to about 99%,from about 80% to about 99% and from about 90% to about 99%. In stillfurther embodiments, the Stevia extract contains Reb X in an amountgreater than about 90% by weight on a dry basis, for example, greaterthan about 91%, greater than about 92%, greater than about 93%, greaterthan about 94%, greater than about 95%, greater than about 96%, greaterthan about 97%, greater than about 98% and greater than about 99%.

In one embodiment, Reb X is provided as a component of a steviolglycoside mixture in a sweetener composition, i.e., a mixture of steviolglycosides wherein the remainder of the non-Reb X portion of the mixtureis comprised entirely of steviol glycosides. The identities of steviolglycosides are known in the art and include, but are not limited to,steviol monoside, rubososide, steviolbioside, stevioside, rebaudiosideA, rebaudioside B, rebaudioside C, rebaudioside D, rebaudioside E,rebaudioside F and dulcoside A. The steviol glycoside mixture maycontain from about 5% to about 99% Reb X by weight on a dry basis. Forexample, a steviol glycoside mixture may contain from about 10% to about99%, from about 20% to about 99%, from about 30% to about 99%, fromabout 40% to about 99%, from about 50% to about 99%, from about 60% toabout 99%, from about 70% to about 99%, from about 80% to about 99% andfrom about 90% to about 99% Reb X by weight on a dry basis. In stillfurther embodiments, the steviol glycoside mixture may contain greaterthan about 90% Reb X by weight on a dry basis, for example, greater thanabout 91%, greater than about 92%, greater than about 93%, greater thanabout 94%, greater than about 95%, greater than about 96%, greater thanabout 97%, greater than about 98% and greater than about 99%.

In one embodiment, Reb X is the sole sweetener in the sweetenercomposition, i.e. Reb X is the only compound present in the sweetenercomposition that provides sweetness. In another embodiment, Reb X is oneof two or more sweetener compounds present in the sweetener composition.

The amount of sucrose in a reference solution may be described indegrees Brix (° Bx). One degree Brix is 1 gram of sucrose in 100 gramsof solution and represents the strength of the solution as percentage byweight (% w/w) (strictly speaking, by mass). In one embodiment, asweetener composition contains Reb X in an amount effective to providesweetness equivalent from about 0.50 to 14 degrees Brix of sugar whenpresent in a sweetened composition, such as, for example, from about 5to about 11 degrees Brix, from about 4 to about 7 degrees Brix, or about5 degrees Brix. In another embodiment, Reb X is present in an amounteffective to provide sweetness equivalent to about 10 degrees Brix whenpresent in a sweetened composition.

The sweetness of a non-sucrose sweetener can also be measured against asucrose reference by determining the non-sucrose sweetener's sucroseequivalence. Typically, taste panelists are trained to detect sweetnessof reference sucrose solutions containing between 1-15% sucrose (w/v).Other non-sucrose sweeteners are then tasted at a series of dilutions todetermine the concentration of the non-sucrose sweetener that is assweet as a given percent sucrose reference. For example, if a 1%solution of a sweetener is as sweet as a 10% sucrose solution, then thesweetener is said to be 10 times as potent as sucrose.

In one embodiment, Reb X is present in an effective amount to provide asucrose equivalence of greater than about 10% (w/v) when present in asweetened composition, such as, for example, greater than about 11%,greater than about 12%, greater than about 13% or greater than about14%.

The amount of Reb X in the sweetener composition may vary. In oneembodiment, Reb X is present in a sweetener composition in any amount toimpart the desired sweetness when the sweetener composition is presentin a sweetened composition. For example, Reb X is present in thesweetener composition in an amount effective to provide a Reb Xconcentration from about 1 ppm to about 10,000 ppm when present in asweetened composition, such as, for example, from about 1 ppm to about4,000 ppm, from about 1 ppm to about 3,000 ppm, from about 1 ppm toabout 2,000 ppm, from about 1 ppm to about 1,000 ppm. In anotherembodiment, Reb X is present in the sweetener composition in an amounteffective to provide a Reb X concentration from about 10 ppm to about1,000 ppm when present in a sweetened composition, such as, for example,from about 10 ppm to about 800 ppm, from about 50 ppm to about 800 ppm,from about 50 ppm to about 600 ppm or from about 200 ppm to about 250ppm. In a particular embodiment, Reb X is present in the sweetenercomposition in an amount effective to provide a Reb X concentration fromabout 300 ppm to about 600 ppm.

In some embodiments, sweetener compositions contain one or moreadditional sweeteners. The additional sweetener can be any type ofsweetener, for example, a natural, non-natural, or synthetic sweetener.In at least one embodiment, the at least one additional sweetener ischosen from natural sweeteners other than Stevia sweeteners. In anotherembodiment, the at least one additional sweetener is chosen fromsynthetic high potency sweeteners.

For example, the at least one additional sweetener may be a carbohydratesweetener. Non-limiting examples of suitable carbohydrate sweetenersinclude sucrose, fructose, glucose, erythritol, maltitol, lactitol,sorbitol, mannitol, xylitol, tagatose, trehalose, galactose, rhamnose,cyclodextrin (e.g., α-cyclodextrin, β-cyclodextrin, and γ-cyclodextrin),ribulose, threose, arabinose, xylose, lyxose, allose, altrose, mannose,idose, lactose, maltose, invert sugar, isotrehalose, neotrehalose,palatinose or isomaltulose, erythrose, deoxyribose, gulose, idose,talose, erythrulose, xylulose, psicose, turanose, cellobiose,glucosamine, mannosamine, fucose, fuculose, glucuronic acid, gluconicacid, glucono-lactone, abequose, galactosamine, xylo-oligosaccharides(xylotriose, xylobiose and the like), gentio-oligoscaccharides(gentiobiose, gentiotriose, gentiotetraosc and the like),galacto-oligosaccharides, sorbose, ketotriose (dehydroxyacetone),aldotriose (glyceraldehyde), nigero-oligosaccharides,fructooligosaccharides (kestose, nystose and the like), maltotetraose,maltotriol, tetrasaccharides, mannan-oligosaccharides,malto-oligosaccharides (maltotriose, maltotetraose, maltopentaose,maltohexaose, maltoheptaose and the like), dextrins, lactulose,melibiose, raffinose, rhamnose, ribose, isomerized liquid sugars such ashigh fructose corn/starch syrup (HFCS/HFSS) (e.g., HFCS55, HFCS42, orHFCS90), coupling sugars, soybean oligosaccharides, glucose syrup andcombinations thereof. D- or L-configurations can be used whenapplicable.

In other embodiments, the additional sweetener is a carbohydratesweetener selected from the group consisting of glucose, fructose,sucrose and combinations thereof.

In another embodiment, the additional sweetener is a carbohydratesweetener selected from D-allose, D-psicose, L-ribose, D-tagatose,L-glucose, L-fucose, L-Arbinose, Turanose and combinations thereof.

The Reb X and carbohydrate sweetener may be present in any weight ratio,such as, for example, from about 0.001:14 to about 1:0.01, such as, forexample, about 0.06:6. Carbohydrates are present in the sweetenercomposition in an amount effective to provide a concentration from about100 ppm to about 140,000 ppm when present in a sweetened composition,such as, for example, a beverage.

In yet other embodiments, the at least one additional sweetener is asynthetic sweetener. As used herein, the phrase “synthetic sweetener”refers to any composition which is not found naturally in nature andcharacteristically has a sweetness potency greater than sucrose,fructose, or glucose, yet has less calories. Non-limiting examples ofsynthetic high-potency sweeteners suitable for embodiments of thisdisclosure include sucralose, potassium acesulfame, acesulfame acid andsalts thereof, aspartame, alitame, saccharin and salts thereof,neohesperidin dihydrochalcone, cyclamate, cyclamic acid and saltsthereof, neotame, advantame, glucosylated steviol glycosides (GSGs) andcombinations thereof. The synthetic sweetener is present in thesweetener composition in an amount effective to provide a concentrationfrom about 0.3 ppm to about 3,500 ppm when present in a sweetenedcomposition, such as, for example, a beverage.

In still other embodiments, the additional sweetener can be a naturalhigh potency sweetener. Suitable natural high potency sweetenersinclude, but are not limited to, rebaudioside A, rebaudioside B,rebaudioside C, rebaudioside D, rebaudioside E, rebaudioside F,rebaudioside I, rebaudioside H, rebaudioside L, rebaudioside K,rebaudioside J, rebaudioside N, rebaudioside O, dulcoside A, dulcosideB, rubusoside, stevia, stevioside, mogroside IV, mogroside V, Luo HanGuo, siamenoside, monatin and its salts (monatin SS, RR, RS, SR),curculin, glycyrrhizic acid and its salts, thaumatin, monellin,mabinlin, brazzein, hernandulcin, phyllodulcin, glycyphyllin,phloridzin, trilobatin, baiyunoside, osladin, polypodoside A,pterocaryoside A, pterocaryoside B, mukurozioside, phlomisoside I,periandrin I, abrusoside A, steviolbioside and cyclocarioside I. Thenatural high potency sweetener can be provided as a pure compound or,alternatively, as part of an extract. For example, rebaudioside A can beprovided as a sole compound or as part of a Stevia extract. The naturalhigh potency sweetener is present in the sweetener composition in anamount effective to provide a concentration from about 0.1 ppm to about3,000 ppm when present in a sweetened composition, such as, for example,a beverage.

In still other embodiments, the additional sweetener can be chemicallyor enzymatically modified natural high potency sweetener. Modifiednatural high potency sweeteners include glycosylated natural highpotency sweetener such as glucosyl-, galactosyl-, fructosyl-derivativescontaining 1-50 glycosidic residues. Glycosylated natural high potencysweeteners may be prepared by enzymatic transglycosylation reactioncatalyzed by various enzymes possessing transglycosylating activity.

In another particular embodiment, a sweetener compositions comprises RebX and at least one other sweetener that functions as the sweetenercomponent (i.e. the substance or substances that provide sweetness) of asweetener composition. The sweetener compositions often exhibit synergywhen combined and have improved flavor and temporal profiles compared toeach sweetener alone. One or more additional sweetener can be used inthe sweetener compositions. In one embodiment, a sweeteners compositioncontains Reb X and one additional sweetener. In other embodiments, asweetener composition contains Reb X and more than one additionalsweetener. The at least one other sweetener can be selected from thegroup consisting of erythritol, Reb B, NSF-02, mogroside V, Reb A, Reb Dand combinations thereof.

In one embodiment, a sweetener composition comprises Reb X anderythritol as the sweetener component. The relative weight percent ofReb X and erythritol can vary. Generally, erythritol can comprise fromabout 0.1% to about 3.5% by weight of the sweetener component.

In another embodiment, a sweetener composition comprises Reb X and Reb Bas the sweetener component. The relative weight percent of Reb X and RebB can each vary from about 1% to about 99%, such as for example, about95% Reb X/5% Reb B, about 90% Reb X/10% Reb B, about 85% Reb X/15% RebB, about 80% Reb X/20% Reb B, about 75% Reb X/25% Reb B, about 70% RebX/30% Reb B, about 65% Reb X/35% Reb B, about 60% Reb X/40% Reb B, about55% Reb X/45% Reb B, about 50% Reb X/50% Reb B, about 45% Reb X/55% RebB, about 40% Reb X/60% Reb B, about 35% Reb X/65% Reb B, about 30% RebX/70% Reb B, about 25% Reb X/75% Reb B, about 20% Reb X/80% Reb B, about15% Reb X/85% Reb B, about 10% Reb X/90% Reb B or about 5% Reb X/10% RebB. In a particular embodiment, Reb B comprises from about 5% to about40% of the sweetener component, such as, for example, from about 10% toabout 30% or about 15% to about 25%.

In yet another embodiment, a sweetener composition comprises Reb X andNSF-02 (a GSG-type sweetener, available from PureCircle) as thesweetener component. The relative weight percent of Reb X and NSF-02 caneach vary from about 1% to about 99%, such as for example, about 95% RebX/5% NSF-02, about 90% Reb X/10% NSF-02, about 85% Reb X/15% NSF-02,about 80% Reb X/20% NSF-02, about 75% Reb X/25% NSF-02, about 70% RebX/30% NSF-02, about 65% Reb X/35% NSF-02, about 60% Reb X/40% NSF-02,about 55% Reb X/45% NSF-02, about 50% Reb X/50% NSF-02, about 45% RebX/55% NSF-02, about 40% Reb X/60% NSF-02, about 35% Reb X/65% NSF-02,about 30% Reb X/70% NSF-02, about 25% Reb X/75% NSF-02, about 20% RebX/80% NSF-02, about 15% Reb X/85% NSF-02, about 10% Reb X/90% NSF-02 orabout 5% Reb X/10% NSF-02. In a particular embodiment, NSF-02 comprisesfrom about 5% to about 50% of the sweetener component, such as, forexample, from about 10% to about 40% or about 30% to about 30%.

In still another embodiment, a sweetener composition comprises Reb X andmogroside V as the sweetener component. The relative weight percent ofReb X and mogroside V can each vary from about 1% to about 99%, such asfor example, about 95% Reb X/5% mogroside V, about 90% Reb X/10%mogroside V, about 85% Reb X/15% mogroside V, about 80% Reb X/20%mogroside V, about 75% Reb X/25% mogroside V, about 70% Reb X/30%mogroside V, about 65% Reb X/35% mogroside V, about 60% Reb X/40%mogroside V, about 55% Reb X/45% mogroside V, about 50% Reb X/50%mogroside V, about 45% Reb X/55% mogroside V, about 40% Reb X/60%mogroside V, about 35% Reb X/65% mogroside V, about 30% Reb X/70%mogroside V, about 25% Reb X/75% mogroside V, about 20% Reb X/80%mogroside V, about 15% Reb X/85% mogroside V, about 10% Reb X/90%mogroside V or about 5% Reb X/10% mogroside V. In a particularembodiment, mogroside V comprises from about 5% to about 50% of thesweetener component, such as, for example, from about 10% to about 40%or about 30% to about 30%.

In another embodiment, a sweetener composition comprises Reb X and Reb Aas the sweetener component. The relative weight percent of Reb X and RebA can each vary from about 1% to about 99%, such as for example, about95% Reb X/5% Reb A, about 90% Reb X/10% Reb A, about 85% Reb X/15% RebA, about 80% Reb X/20% Reb A, about 75% Reb X/25% Reb A, about 70% RebX/30% Reb A, about 65% Reb X/35% Reb A, about 60% Reb X/40% Reb A, about55% Reb X/45% Reb A, about 50% Reb X/50% Reb A, about 45% Reb X/55% RebA, about 40% Reb X/60% Reb A, about 35% Reb X/65% Reb A, about 30% RebX/70% Reb A, about 25% Reb X/75% Reb A, about 20% Reb X/80% Reb A, about15% Reb X/85% Reb A, about 10% Reb X/90% Reb A or about 5% Reb X/10% RebA. In a particular embodiment, Reb A comprises from about 5% to about40% of the sweetener component, such as, for example, from about 10% toabout 30% or about 15% to about 25%.

In another embodiment, a sweetener composition comprises Reb X and Reb Das the sweetener component. The relative weight percent of Reb X and RebD can each vary from about 1% to about 99%, such as for example, about95% Reb X/5% Reb D, about 90% Reb X/10% Reb D, about 85% Reb X/15% RebD, about 80% Reb X/20% Reb D, about 75% Reb X/25% Reb D, about 70% RebX/30% Reb D, about 65% Reb X/35% Reb D, about 60% Reb X/40% Reb D, about55% Reb X/45% Reb D, about 50% Reb X/50% Reb D, about 45% Reb X/55% RebD, about 40% Reb X/60% Reb D, about 35% Reb X/65% Reb D, about 30% RebX/70% Reb D, about 25% Reb X/75% Reb D, about 20% Reb X/80% Reb D, about15% Reb X/85% Reb D, about 10% Reb X/90% Reb D or about 5% Reb X/10% RebD. In a particular embodiment, Reb D comprises from about 5% to about40% of the sweetener component, such as, for example, from about 10% toabout 30% or about 15% to about 25%.

In another embodiment, a sweetener composition comprises Reb X, Reb Aand Reb D as the sweetener component. The relative weight percent of RebX, Reb D and Reb A can each vary from about 1% to about 99%.

In still another embodiment, a sweetener composition comprises Reb X,Reb B and Reb D as the sweetener component. The relative weight percentof Reb X, Reb B and Reb D can each vary from about 1% to about 99%.

The sweetener compositions can be customized to provide the desiredcalorie content. For example, sweetener compositions can be“full-calorie”, such that they impart the desired sweetness when addedto a sweetenable composition (such as, for example, a beverage) and haveabout 120 calories per 8 oz serving. Alternatively, sweetenercompositions can be “mid-calorie”, such that they impart the desiredsweetness when added to a sweetenable composition (such as, for example,as beverage) and have less than about 60 calories per 8 oz serving. Inother embodiments, sweetener compositions can be “low-calorie”, suchthat they impart the desired sweetness when added to a sweetenablecomposition (such as, for example, as beverage) and have less than 40calories per 8 oz serving. In still other embodiments, the sweetenercompositions can be “zero-calorie”, such that they impart the desiredsweetness when added to a sweetenable composition (such as, for example,a beverage) and have less than 5 calories per 8 oz. serving.

Additives

In addition to Reb X and, optionally, other sweeteners, the sweetenercompositions can optionally include additional additives, detailedherein below. In some embodiments, the sweetener composition containsadditives including, but not limited to, carbohydrates, polyols, aminoacids and their corresponding salts, poly-amino acids and theircorresponding salts, sugar acids and their corresponding salts,nucleotides, organic acids, inorganic acids, organic salts includingorganic acid salts and organic base salts, inorganic salts, bittercompounds, flavorants and flavoring ingredients, astringent compounds,proteins or protein hydrolysates, surfactants, emulsifiers, weighingagents, gums, antioxidants, colorants, flavonoids, alcohols, polymersand combinations thereof. In some embodiments, the additives act toimprove the temporal and flavor profile of the sweetener to provide asweetener composition with a taste similar to sucrose.

In one embodiment, the sweetener compositions contain one or morepolyols. The term “polyol”, as used herein, refers to a molecule thatcontains more than one hydroxyl group. A polyol may be a diol, triol, ora tetraol which contains 2, 3, and 4 hydroxyl groups respectively. Apolyol also may contain more than 4 hydroxyl groups, such as a pentaol,hexaol, heptaol, or the like, which contain 5, 6, or 7 hydroxyl groups,respectively. Additionally, a polyol also may be a sugar alcohol,polyhydric alcohol, or polyalcohol which is a reduced form ofcarbohydrate, wherein the carbonyl group (aldehyde or ketone, reducingsugar) has been reduced to a primary or secondary hydroxyl group.

Non-limiting examples of polyols in some embodiments include erythritol,maltitol, mannitol, sorbitol, lactitol, xylitol, isomalt, propyleneglycol, glycerol (glycerin), threitol, galactitol, palatinose, reducedisomalto-oligosaccharides, reduced xylo-oligosaccharides, reducedgentio-oligosaccharides, reduced maltose syrup, reduced glucose syrup,and sugar alcohols or any other carbohydrates capable of being reducedwhich do not adversely affect the taste of the sweetener composition.

In certain embodiments, the polyol is present in the sweetenercomposition in an amount effective to provide a concentration from about100 ppm to about 250,000 ppm when present in a sweetened composition,such as, for example, a beverage. In other embodiments, the polyol ispresent in the sweetener composition in an amount effective to provide aconcentration from about 400 ppm to about 80,000 ppm when present in asweetened composition, such as, for example, from about 5,000 ppm toabout 40,000 ppm.

In other embodiments, Reb X and the polyol are present in the sweetenercomposition in a weight ratio from about 1:1 to about 1:800, such as,for example, from about 1:4 to about 1:800, from about 1:20 to about1:600, from about 1:50 to about 1:300 or from about 1:75 to about 1:150.

Suitable amino acid additives include, but are not limited to, asparticacid, arginine, glycine, glutamic acid, proline, threonine, theanine,cysteine, cystine, alanine, valine, tyrosine, leucine, arabinose,trans-4-hydroxyproline, isoleucine, asparagine, serine, lysine,histidine, ornithine, methionine, carnitine, aminobutyric acid (α-, β-,and/or δ-isomers), glutamine, hydroxyproline, taurine, norvaline,sarcosine, and their salt forms such as sodium or potassium salts oracid salts. The amino acid additives also may be in the D- orL-configuration and in the mono-, di-, or tri-form of the same ordifferent amino acids. Additionally, the amino acids may be α-, β-, γ-and/or δ-isomers if appropriate. Combinations of the foregoing aminoacids and their corresponding salts (e.g., sodium, potassium, calcium,magnesium salts or other alkali or alkaline earth metal salts thereof,or acid salts) also are suitable additives in some embodiments. Theamino acids may be natural or synthetic. The amino acids also may bemodified. Modified amino acids refers to any amino acid wherein at leastone atom has been added, removed, substituted, or combinations thereof(e.g., N-alkyl amino acid, N-acyl amino acid, or N-methyl amino acid).Non-limiting examples of modified amino acids include amino acidderivatives such as trimethyl glycine, N-methyl-glycine, andN-methyl-alanine. As used herein, modified amino acids encompass bothmodified and unmodified amino acids. As used herein, amino acids alsoencompass both peptides and polypeptides (e.g., dipeptides, tripeptides,tetrapeptides, and pentapeptides) such as glutathione andL-alanyl-L-glutamine. Suitable polyamino acid additives includepoly-L-aspartic acid, poly-L-lysine (e.g., poly-L-α-lysine orpoly-L-ε-lysine), poly-L-ornithine (e.g., poly-L-α-ornithine orpoly-L-ε-ornithine), poly-L-arginine, other polymeric forms of aminoacids, and salt forms thereof (e.g., calcium, potassium, sodium, ormagnesium salts such as L-glutamic acid mono sodium salt). Thepoly-amino acid additives also may be in the D- or L-configuration.Additionally, the poly-amino acids may be α-, β-, γ-, δ-, and ε-isomersif appropriate. Combinations of the foregoing poly-amino acids and theircorresponding salts (e.g., sodium, potassium, calcium, magnesium saltsor other alkali or alkaline earth metal salts thereof or acid salts)also are suitable additives in some embodiments. The poly-amino acidsdescribed herein also may comprise co-polymers of different amino acids.The poly-amino acids may be natural or synthetic. The poly-amino acidsalso may be modified, such that at least one atom has been added,removed, substituted, or combinations thereof (e.g., N-alkyl poly-aminoacid or N-acyl poly-amino acid). As used herein, poly-amino acidsencompass both modified and unmodified poly-amino acids. For example,modified poly-amino acids include, but are not limited to, poly-aminoacids of various molecular weights (MW), such as poly-L-α-lysine with aMW of 1,500, MW of 6,000, MW of 25,200, MW of 63,000, MW of 83,000, orMW of 300,000.

In particular embodiments, the amino acid is present in the sweetenercomposition in an amount effective to provide a concentration from about10 ppm to about 50,000 ppm when present in a sweetened composition, suchas, for example, a beverage. In another embodiment, the amino acid ispresent in the sweetener composition in an amount effective to provide aconcentration from about 1,000 ppm to about 10,000 ppm when present in asweetened composition, such as, for example, from about 2,500 ppm toabout 5,000 ppm or from about 250 ppm to about 7,500 ppm.

Suitable sugar acid additives include, but are not limited to, aldonic,uronic, aldaric, alginic, gluconic, glucuronic, glucaric, galactaric,galacturonic, and salts thereof (e.g., sodium, potassium, calcium,magnesium salts or other physiologically acceptable salts), andcombinations thereof.

Suitable nucleotide additives include, but are not limited to, inosinemonophosphate (“IMP”), guanosine monophosphate (“GMP”), adenosinemonophosphate (“AMP”), cytosine monophosphate (CMP), uracilmonophosphate (UMP), inosine diphosphate, guanosine diphosphate,adenosine diphosphate, cytosine diphosphate, uracil diphosphate, inosinetriphosphate, guanosine triphosphate, adenosine triphosphate, cytosinetriphosphate, uracil triphosphate, alkali or alkaline earth metal saltsthereof, and combinations thereof. The nucleotides described herein alsomay comprise nucleotide-related additives, such as nucleosides ornucleic acid bases (e.g., guanine, cytosine, adenine, thymine, uracil).

The nucleotide is present in the sweetener composition in an amounteffective to provide a concentration from about 5 ppm to about 1,000 ppmwhen present in sweetened composition, such as, for example, a beverage.

Suitable organic acid additives include any compound which comprises a—COOH moiety, such as, for example, C2-C30 carboxylic acids, substitutedhydroxyl C2-C30 carboxylic acids, butyric acid (ethyl esters),substituted butyric acid (ethyl esters), benzoic acid, substitutedbenzoic acids (e.g., 2,4-dihydroxybenzoic acid), substituted cinnamicacids, hydroxyacids, substituted hydroxybenzoic acids, anisic acidsubstituted cyclohexyl carboxylic acids, tannic acid, aconitic acid,lactic acid, tartaric acid, citric acid, isocitric acid, gluconic acid,glucoheptonic acids, adipic acid, hydroxycitric acid, malic acid,fruitaric acid (a blend of malic, fumaric, and tartaric acids), fumaricacid, maleic acid, succinic acid, chlorogenic acid, salicylic acid,creatine, caffeic acid, bile acids, acetic acid, ascorbic acid, alginicacid, erythorbic acid, polyglutamic acid, glucono delta lactone, andtheir alkali or alkaline earth metal salt derivatives thereof. Inaddition, the organic acid additives also may be in either the D- orL-configuration.

Suitable organic acid additive salts include, but are not limited to,sodium, calcium, potassium, and magnesium salts of all organic acids,such as salts of citric acid, malic acid, tartaric acid, fumaric acid,lactic acid (e.g., sodium lactate), alginic acid (e.g., sodiumalginate), ascorbic acid (e.g., sodium ascorbate), benzoic acid (e.g.,sodium benzoate or potassium benzoate), sorbic acid and adipic acid. Theexamples of the organic acid additives described optionally may besubstituted with at least one group chosen from hydrogen, alkyl,alkenyl, alkynyl, halo, haloalkyl, carboxyl, acyl, acyloxy, amino,amido, carboxyl derivatives, alkylamino, dialkylamino, acylamino,alkoxy, aryloxy, nitro, cyano, sulfo, thiol, imine, sulfonyl, sulfonyl,sulfinyl, sulfamyl, carboxalkoxy, carboxamido, phosphonyl, phosphinyl,phosphoryl, phosphino, thioester, thioether, anhydride, oximino,hydrazino, carbamyl, phosphor or phosphonato. In particular embodiments,the organic acid additive is present in the sweetener composition in anamount from about 10 ppm to about 5,000 ppm.

Suitable inorganic acid additives include, but are not limited to,phosphoric acid, phosphorous acid, polyphosphoric acid, hydrochloricacid, sulfuric acid, carbonic acid, sodium dihydrogen phosphate, andalkali or alkaline earth metal salts thereof (e.g., inositolhexaphosphate Mg/Ca).

The inorganic acid additive is present in the sweetener composition inan amount effective to provide a concentration from about 25 ppm toabout 25,000 ppm when present in a sweetened composition, such as, forexample, a beverage.

Suitable bitter compound additives include, but are not limited to,caffeine, quinine, urea, bitter orange oil, naringin, quassia, and saltsthereof.

The bitter compound is present in the sweetener composition in an amounteffective to provide a concentration from about 25 ppm to about 25,000ppm when present in a sweetened composition, such as, for example, abeverage.

Suitable flavorant and flavoring ingredient additives for include, butare not limited to, vanillin, vanilla extract, mango extract, cinnamon,citrus, coconut, ginger, viridiflorol, almond, menthol (includingmenthol without mint), grape skin extract, and grape seed extract.“Flavorant” and “flavoring ingredient” are synonymous and can includenatural or synthetic substances or combinations thereof. Flavorants alsoinclude any other substance which imparts flavor and may include naturalor non-natural (synthetic) substances which are safe for human oranimals when used in a generally accepted range. Non-limiting examplesof proprietary flavorants include Döhler™ Natural Flavoring SweetnessEnhancer K14323 (Döhler™, Darmstadt, Germany), Symrise™ Natural FlavorMask for Sweeteners 161453 and 164126 (Symrise™, Holzminden, Germany),Natural Advantage™ Bitterness Blockers 1, 2, 9 and 10 (NaturalAdvantage™, Freehold, N.J., U.S.A.), and Sucramask™ (Creative ResearchManagement, Stockton, Calif., U.S.A.).

The flavorant is present in the sweetener composition in an amounteffective to provide a concentration from about 0.1 ppm to about 4,000ppm when present in a sweetened composition, such as, for example, abeverage.

Suitable polymer additives include, but are not limited to, chitosan,pectin, pectic, pectinic, polyuronic, polygalacturonic acid, starch,food hydrocolloid or crude extracts thereof (e.g., gum acacia senegal(Fibergum™), gum acacia seyal, carageenan), poly-L-lysine (e.g.,poly-L-α-lysine or poly-L-ε-lysine), poly-L-ornithine (e.g.,poly-L-α-ornithine or poly-L-ε-ornithine), polypropylene glycol,polyethylene glycol, poly(ethylene glycol methyl ether), polyarginine,polyaspartic acid, polyglutamic acid, polyethylene imine, alginic acid,sodium alginate, propylene glycol alginate, and sodiumpolyethyleneglycolalginate, sodium hexametaphosphate and its salts, andother cationic polymers and anionic polymers.

The polymer is present in the sweetener composition in an amounteffective to provide a concentration from about 30 ppm to about 2,000ppm when present in a sweetened composition, such as, for example, abeverage.

Suitable protein or protein hydrolysate additives include, but are notlimited to, bovine serum albumin (BSA), whey protein (includingfractions or concentrates thereof such as 90% instant whey proteinisolate, 34% whey protein, 50% hydrolyzed whey protein, and 80% wheyprotein concentrate), soluble rice protein, soy protein, proteinisolates, protein hydrolysates, reaction products of proteinhydrolysates, glycoproteins, and/or proteoglycans containing amino acids(e.g., glycine, alanine, serine, threonine, asparagine, glutamine,arginine, valine, isoleucine, leucine, norvaline, methionine, proline,tyrosine, hydroxyproline, and the like), collagen (e.g., gelatin),partially hydrolyzed collagen (e.g., hydrolyzed fish collagen), andcollagen hydrolysates (e.g., porcine collagen hydrolysate).

The protein hydrosylate is present in the sweetener composition in anamount effective to provide a concentration from about 200 ppm to about50,000 ppm when present in a sweetened composition, such as, forexample, a beverage.

Suitable surfactant additives include, but are not limited to,polysorbates (e.g., polyoxyethylene sorbitan monooleate (polysorbate80), polysorbate 20, polysorbate 60), sodium dodecylbenzenesulfonate,dioctyl sulfosuccinate or dioctyl sulfosuccinate sodium, sodium dodecylsulfate, cetylpyridinium chloride (hexadecylpyridinium chloride),hexadecyltrimethylammonium bromide, sodium cholate, carbamoyl, cholinechloride, sodium glycocholate, sodium taurodeoxycholate, lauricarginate, sodium stearoyl lactylate, sodium taurocholate, lecithins,sucrose oleate esters, sucrose stearate esters, sucrose palmitateesters, sucrose laurate esters, and other emulsifiers, and the like.

The surfactant additive is present in the sweetener composition in anamount effective to provide a concentration from about 30 ppm to about2,000 ppm when present in a sweetened composition, such as, for example,a beverage.

Suitable flavonoid additives are classified as flavonols, flavones,flavanones, flavan-3-ols, isoflavones, or anthocyanidins. Non-limitingexamples of flavonoid additives include, but are not limited to,catechins (e.g., green tea extracts such as Polyphenon™ 60, Polyphenon™30, and Polyphenon™ 25 (Mitsui Norin Co., Ltd., Japan), polyphenols,rutins (e.g., enzyme modified rutin Sanmelin™ AO (San-fi Gen F.F.I.,Inc., Osaka, Japan)), neohesperidin, naringin, neohesperidindihydrochalcone, and the like.

The flavonoid additive is present in the sweetener composition in anamount effective to provide a concentration from about 0.1 ppm to about1,000 ppm when present in sweetened composition, such as, for example, abeverage.

Suitable alcohol additives include, but are not limited to, ethanol. Inparticular embodiments, the alcohol additive is present in the sweetenercomposition in an amount effective to provide a concentration from about625 ppm to about 10,000 ppm when present in a sweetened composition,such as, for example, a beverage.

Suitable astringent compound additives include, but are not limited to,tannic acid, europium chloride (EuCl₃), gadolinium chloride (GdCl₃),terbium chloride (TbCl₃), alum, tannic acid, and polyphenols (e.g., teapolyphenols). The astringent additive is present in the sweetenercomposition in an amount effective to provide a concentration from about10 ppm to about 5,000 ppm when present in a sweetened composition, suchas, for example, a beverage.

In particular embodiments, a sweetener composition comprises Reb X; apolyol selected from erythritol, maltitol, mannitol, xylitol, sorbitol,and combinations thereof and optionally at least one additionalsweetener and/or functional ingredient. The Reb X can be provided as apure compound or as part of a Stevia extract or steviol glycosidemixture, as described above. Reb X can be present in an amount fromabout 5% to about 99% by weight on a dry basis in either a steviolglycoside mixture or a Stevia extract. In one embodiment, Reb X and thepolyol are present in a sweetener composition in a weight ratio fromabout 1:1 to about 1:800, such as, for example, from about 1:4 to about1:800, from about 1:20 to about 1:600, from about 1:50 to about 1:300 orfrom about 1:75 to about 1:150. In another embodiment, Reb X is presentin the sweetener composition in an amount effective to provide aconcentration from about 1 ppm to about 10,000 ppm when present in asweetened composition, such as, for example, about 300 ppm. The polyol,such as, for example, erythritol, can be present in the sweetenercomposition in an amount effective to provide a concentration from about100 ppm to about 250,000 ppm when present in a sweetened composition,such as, for example, from about 5,000 ppm to about 40,000 ppm, fromabout 1,000 ppm to about 35,000 ppm.

In particular embodiments, a sweetener composition comprises Reb X; acarbohydrate sweetener selected from sucrose, fructose, glucose, maltoseand combinations thereof; and optionally at least one additionalsweetener and/or functional ingredient. The Reb X can be provided as apure compound or as part of a Stevia extract or steviol glycosidemixture, as described above. Reb X can be present in an amount fromabout 5% to about 99% by weight on a dry basis in either a steviolglycoside mixture or a Stevia extract. In one embodiment, Reb X and thecarbohydrate are present in a sweetener composition in a weight ratiofrom about 0.001:14 to about 1:0.01, such as, for example, about 0.06:6.In one embodiment, Reb X is present in the sweetener composition in anamount effective to provide a concentration from about 1 ppm to about10,000 ppm when present in a sweetened composition, such as, forexample, about 500 ppm. The carbohydrate, such as, for example, sucrose,can be present in the sweetener composition in an amount effective toprovide a concentration from about 100 ppm to about 140,000 ppm whenpresent in a sweetened composition, such as, for example, from about1,000 ppm to about 100,000 ppm, from about 5,000 ppm to about 80,000ppm.

In particular embodiments, a sweetener composition comprises Reb X; anamino acid selected from glycine, alanine, proline and combinationsthereof; and optionally at least one additional sweetener and/orfunctional ingredient. The Reb X can be provided as a pure compound oras part of a Stevia extract or steviol glycoside mixture, as describedabove. Reb X can be present in an amount from about 5% to about 99% byweight on a dry basis in either a steviol glycoside mixture or a Steviaextract. In another embodiment, Reb X is present in the sweetenercomposition in an amount effective to provide a concentration from about1 ppm to about 10,000 ppm when present in a sweetened composition, suchas, for example, about 500 ppm. The amino acid, such as, for example,glycine, can be present in the sweetener composition in an amounteffective to provide a concentration from about 10 ppm to about 50,000ppm when present in a sweetened composition, such as, for example, fromabout 1,000 ppm to about 10,000 ppm, from about 2,500 ppm to about 5,000ppm

In particular embodiments, a sweetener composition comprises Reb X; asalt selected from sodium chloride, magnesium chloride, potassiumchloride, calcium chloride and combinations thereof; and optionally atleast one additional sweetener and/or functional ingredient. The Reb Xcan be provided as a pure compound or as part of a Stevia extract orsteviol glycoside mixture, as described above. Reb X can be present inan amount from about 5% to about 99% by weight on a dry basis in eithera steviol glycoside mixture or a Stevia extract. In one embodiment, RebX is present in the sweetener composition in an amount effective toprovide a concentration from about 1 ppm to about 10,000 ppm, such as,for example, about 100 to about 1,000 ppm. The inorganic salt, such as,for example, magnesium chloride, is present in the sweetener compositionin an amount effective to provide a concentration from about 25 ppm toabout 25,000 ppm when present in a sweetened composition, such as, forexample, from about 100 ppm to about 4,000 ppm or from about 100 ppm toabout 3,000 ppm.

Functional Ingredients

The sweetener composition can also contain one or more functionalingredients, which provide a real or perceived heath benefit to thecomposition. Functional ingredients include, but are not limited to,saponins, antioxidants, dietary fiber sources, fatty acids, vitamins,glucosamine, minerals, preservatives, hydration agents, probiotics,prebiotics, weight management agents, osteoporosis management agents,phytoestrogens, long chain primary aliphatic saturated alcohols,phytosterols and combinations thereof.

Saponin

In certain embodiments, the functional ingredient is at least onesaponin. In one embodiment, a sweetener composition comprises at leastone saponin, Reb X, and optionally at least one additive. In anotherembodiment, a sweetened composition comprises a sweetenable composition,at least one saponin, Reb X, and optionally at least one additive. Instill another embodiment, a sweetened composition comprises asweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one saponin, Reb X, andoptionally at least one additive.

As used herein, the at least one saponin may comprise a single saponinor a plurality of saponins as a functional ingredient for the sweetenercomposition or sweetened compositions provided herein. Generally,according to particular embodiments of this invention, the at least onesaponin is present in the sweetener composition or sweetened compositionin an amount sufficient to promote health and wellness.

Saponins are glycosidic natural plant products comprising an aglyconering structure and one or more sugar moieties. The combination of thenonpolar aglycone and the water soluble sugar moiety gives saponinssurfactant properties, which allow them to form a foam when shaken in anaqueous solution.

The saponins are grouped together based on several common properties. Inparticular, saponins are surfactants which display hemolytic activityand form complexes with cholesterol. Although saponins share theseproperties, they are structurally diverse. The types of aglycone ringstructures forming the ring structure in saponins can vary greatly.Non-limiting examples of the types of aglycone ring structures insaponin for use in particular embodiments of the invention includesteroids, triterpenoids, and steroidal alkaloids. Non-limiting examplesof specific aglycone ring structures for use in particular embodimentsof the invention include soyasapogenol A, soyasapogenol B andsoyasopogenol E. The number and type of sugar moieties attached to theaglycone ring structure can also vary greatly. Non-limiting examples ofsugar moieties for use in particular embodiments of the inventioninclude glucose, galactose, glucuronic acid, xylose, rhamnose, andmethylpentose moieties. Non-limiting examples of specific saponins foruse in particular embodiments of the invention include group A acetylsaponin, group B acetyl saponin, and group E acetyl saponin.

Saponins can be found in a large variety of plants and plant products,and are especially prevalent in plant skins and barks where they form awaxy protective coating. Several common sources of saponins includesoybeans, which have approximately 5% saponin content by dry weight,soapwort plants (Saponaria), the root of which was used historically assoap, as well as alfalfa, aloe, asparagus, grapes, chickpeas, yucca, andvarious other beans and weeds. Saponins may be obtained from thesesources by using extraction techniques well known to those of ordinaryskill in the art. A description of conventional extraction techniquescan be found in U.S. Pat. Appl. No. 2005/0123662, the disclosure ofwhich is expressly incorporated by reference.

Antioxidant

In certain embodiments, the functional ingredient is at least oneantioxidant. In one embodiment, a sweetener composition comprises atleast one antioxidant, Reb X, and optionally at least one additive. Inanother embodiment, a sweetened composition comprises a sweetenablecomposition, at least one antioxidant, Reb X, and optionally at leastone additive. In still another embodiment, a sweetened compositioncomprises a sweetenable composition and a sweetener composition, whereinthe sweetener composition comprises at least one antioxidant, Reb X, andoptionally, at least one additive.

As used herein, the at least one antioxidant may comprise a singleantioxidant or a plurality of antioxidants as a functional ingredientfor the sweetener composition or sweetened compositions provided herein.Generally, according to particular embodiments of this invention, the atleast one antioxidant is present in the sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

As used herein “antioxidant” refers to any substance which inhibits,suppresses, or reduces oxidative damage to cells and biomolecules.Without being bound by theory, it is believed that antioxidants inhibit,suppress, or reduce oxidative damage to cells or biomolecules bystabilizing free radicals before they can cause harmful reactions. Assuch, antioxidants may prevent or postpone the onset of somedegenerative diseases.

Examples of suitable antioxidants for embodiments of this inventioninclude, but are not limited to, vitamins, vitamin cofactors, minerals,hormones, carotenoids, carotenoid terpenoids, non-carotenoid terpenoids,flavonoids, flavonoid polyphenolics (e.g., bioflavonoids), flavonols,flavones, phenols, polyphenols, esters of phenols, esters ofpolyphenols, nonflavonoid phenolics, isothiocyanates, and combinationsthereof. In some embodiments, the antioxidant is vitamin A, vitamin C,vitamin E, ubiquinone, mineral selenium, manganese, melatonin,α-carotene, β-carotene, lycopene, lutein, zeanthin, crypoxanthin,reservatol, eugenol, quercetin, catechin, gossypol, hesperetin,curcumin, ferulic acid, thymol, hydroxytyrosol, tumeric, thyme, oliveoil, lipoic acid, glutathinone, gutamine, oxalic acid,tocopherol-derived compounds, butylated hydroxyanisole (BHA), butylatedhydroxytoluene (BHT), ethylenediaminetetraacetic acid (EDTA),tert-butylhydroquinone, acetic acid, pectin, tocotrienol, tocopherol,coenzyme Q10, zeaxanthin, astaxanthin, canthaxantin, saponins,limonoids, kaempfedrol, myricetin, isorhamnetin, proanthocyanidins,quercetin, rutin, luteolin, apigenin, tangeritin, hesperetin,naringenin, erodictyol, flavan-3-ols (e.g., anthocyanidins),gallocatechins, epicatechin and its gallate forms, epigallocatechin andits gallate forms (ECGC) theaflavin and its gallate forms, thearubigins,isoflavone phytoestrogens, genistcin, daidzein, glycitein,anythocyanins, cyaniding, delphinidin, malvidin, pelargonidin, peonidin,petunidin, ellagic acid, gallic acid, salicylic acid, rosmarinic acid,cinnamic acid and its derivatives (e.g., ferulic acid), chlorogenicacid, chicoric acid, gallotannins, ellagitannins, anthoxanthins,betacyanins and other plant pigments, silymarin, citric acid, lignan,antinutrients, bilirubin, uric acid, R-α-lipoic acid, N-acetylcysteine,emblicanin, apple extract, apple skin extract (applephenon), rooibosextract red, rooibos extract, green, hawthorn berry extract, redraspberry extract, green coffee antioxidant (GCA), aronia extract 20%,grape seed extract (VinOseed), cocoa extract, hops extract, mangosteenextract, mangosteen hull extract, cranberry extract, pomegranateextract, pomegranate hull extract, pomegranate seed extract, hawthornberry extract, pomella pomegranate extract, cinnamon bark extract, grapeskin extract, bilberry extract, pine bark extract, pycnogenol,elderberry extract, mulberry root extract, wolfberry (gogi) extract,blackberry extract, blueberry extract, blueberry leaf extract, raspberryextract, turmeric extract, citrus bioflavonoids, black currant, ginger,acai powder, green coffee bean extract, green tea extract, and phyticacid, or combinations thereof. In alternate embodiments, the antioxidantis a synthetic antioxidant such as butylated hydroxytolune or butylatedhydroxyanisole, for example. Other sources of suitable antioxidants forembodiments of this invention include, but are not limited to, fruits,vegetables, tea, cocoa, chocolate, spices, herbs, rice, organ meats fromlivestock, yeast, whole grains, or cereal grains.

Particular antioxidants belong to the class of phytonutrients calledpolyphenols (also known as “polyphenolics”), which are a group ofchemical substances found in plants, characterized by the presence ofmore than one phenol group per molecule. A variety of health benefitsmay be derived from polyphenols, including prevention of cancer, heartdisease, and chronic inflammatory disease and improved mental strengthand physical strength, for example. Suitable polyphenols for embodimentsof this invention, include catechins, proanthocyanidins, procyanidins,anthocyanins, quercerin, rutin, reservatrol, isoflavones, curcumin,punicalagin, ellagitannin, hesperidin, naringin, citrus flavonoids,chlorogenic acid, other similar materials, and combinations thereof.

In particular embodiments, the antioxidant is a catechin such as, forexample, epigallocatechin gallate (EGCG). Suitable sources of catechinsfor embodiments of this invention include, but arc not limited to, greentea, white tea, black tea, oolong tea, chocolate, cocoa, red wine, grapeseed, red grape skin, purple grape skin, red grape juice, purple grapejuice, berries, pycnogenol, and red apple peel.

In some embodiments, the antioxidant is chosen from proanthocyanidins,procyanidins or combinations thereof. Suitable sources ofproanthocyanidins and procyanidins for embodiments of this inventioninclude, but are not limited to, red grapes, purple grapes, cocoa,chocolate, grape seeds, red wine, cacao beans, cranberry, apple peel,plum, blueberry, black currants, choke berry, green tea, sorghum,cinnamon, barley, red kidney bean, pinto bean, hops, almonds, hazelnuts,pecans, pistachio, pycnogenol, and colorful berries.

In particular embodiments, the antioxidant is an anthocyanin. Suitablesources of anthocyanins for embodiments of this invention include, butare not limited to, red berries, blueberries, bilberry, cranberry,raspberry, cherry, pomegranate, strawberry, elderberry, choke berry, redgrape skin, purple grape skin, grape seed, red wine, black currant, redcurrant, cocoa, plum, apple peel, peach, red pear, red cabbage, redonion, red orange, and blackberries.

In some embodiments, the antioxidant is chosen from quercetin, rutin orcombinations thereof. Suitable sources of quercetin and rutin forembodiments of this invention include, but are not limited to, redapples, onions, kale, bog whortleberry, lingonberrys, chokebeny,cranberry, blackberry, blueberry, strawberry, raspberry, black currant,green tea, black tea, plum, apricot, parsley, leek, broccoli, chilipepper, berry wine, and ginkgo.

In some embodiments, the antioxidant is resveratrol. Suitable sources ofresveratrol for embodiments of this invention include, but are notlimited to, red grapes, peanuts, cranberry, blueberry, bilberry,mulberry, Japanese Itadori tea, and red wine.

In particular embodiments, the antioxidant is an isoflavone. Suitablesources of isoflavones for embodiments of this invention include, butare not limited to, soy beans, soy products, legumes, alfalfa spouts,chickpeas, peanuts, and red clover.

In some embodiments, the antioxidant is curcumin. Suitable sources ofcurcumin for embodiments of this invention include, but are not limitedto, turmeric and mustard.

In particular embodiments, the antioxidant is chosen from punicalagin,ellagitannin or combinations thereof. Suitable sources of punicalaginand ellagitannin for embodiments of this invention include, but are notlimited to, pomegranate, raspberry, strawberry, walnut, and oak-aged redwine.

In some embodiments, the antioxidant is a citrus flavonoid, such ashesperidin or naringin. Suitable sources of citrus flavonids, such ashesperidin or naringin, for embodiments of this invention include, butarc not limited to, oranges, grapefruits, and citrus juices.

In particular embodiments, the antioxidant is chlorogenic acid. Suitablesources of chlorogenic acid for embodiments of this invention include,but are not limited to, green coffee, yerba mate, red wine, grape seed,red grape skin, purple grape skin, red grape juice, purple grape juice,apple juice, cranberry, pomegranate, blueberry, strawberry, sunflower,Echinacea, pycnogenol, and apple peel.

Dietary Fiber

In certain embodiments, the functional ingredient is at least onedietary fiber source. In one embodiment, a sweetener compositioncomprises at least one dietary fiber source, Reb X, and optionally atleast one additive. In another embodiment, a sweetened compositioncomprises a sweetenable composition, at least one dietary fiber source,Reb X, and optionally at least one additive. In still anotherembodiment, a sweetened composition comprises a sweetenable compositionand a sweetener composition, wherein the sweetener composition comprisesat least one dietary fiber source, Reb X, and optionally at least oneadditive.

As used herein, the at least one dietary fiber source may comprise asingle dietary fiber source or a plurality of dietary fiber sources as afunctional ingredient for the sweetener compositions or sweetenedcompositions provided herein. Generally, according to particularembodiments of this invention, the at least one dietary fiber source ispresent in the sweetener composition or sweetened composition in anamount sufficient to promote health and wellness.

Numerous polymeric carbohydrates having significantly differentstructures in both composition and linkages fall within the definitionof dietary fiber. Such compounds are well known to those skilled in theart, non-limiting examples of which include non-starch polysaccharides,lignin, cellulose, methylcellulose, the hemicelluloses, β-glucans,pectins, gums, mucilage, waxes, inulins, oligosaccharides,fructooligosaccharides, cyclodextrins, chitins, and combinationsthereof.

Polysaccharides are complex carbohydrates composed of monosaccharidesjoined by glycosidic linkages. Non-starch polysaccharides are bondedwith β-linkages, which humans are unable to digest due to a lack of anenzyme to break the β-linkages. Conversely, digestable starchpolysaccharides generally comprise α(1-4) linkages.

Lignin is a large, highly branched and cross-linked polymer based onoxygenated phenylpropane units. Cellulose is a linear polymer of glucosemolecules joined by a β(1-4) linkage, which mammalian amylases arcunable to hydrolyze. Methylcellulose is a methyl esther of cellulosethat is often used in foodstuffs as a thickener, and emulsifier. It iscommercially available (e.g., Citrucel by GlaxoSmithKline, Celevac byShire Pharmaceuticals). Hemicelluloses arc highly branched polymersconsisting mainly of glucurono- and 4-O-methylglucuroxylans. β-Glucansare mixed-linkage (1-3), (1-4) β-D-glucose polymers found primarily incereals, such as oats and barley. Pectins, such as beta pectin, are agroup of polysaccharides composed primarily of D-galacturonic acid,which is methoxylated to variable degrees.

Gums and mucilages represent a broad array of different branchedstructures. Guar gum, derived from the ground endosperm of the guarseed, is a galactomannan. Guar gum is commercially available (e.g.,Benefiber by Novartis AG). Other gums, such as gum arabic and pectins,have still different structures. Still other gums include xanthan gum,gellan gum, tara gum, psylium seed husk gum, and locust been gum.

Waxes are esters of ethylene glycol and two fatty acids, generallyoccurring as a hydrophobic liquid that is insoluble in water.

Inulins comprise naturally occurring oligosaccharides belonging to aclass of carbohydrates known as fructans. They generally are comprisedof fructose units joined by β(2-1) glycosidic linkages with a terminalglucose unit. Oligosaccharides are saccharide polymers containingtypically three to six component sugars. They are generally found eitherO- or N-linked to compatible amino acid side chains in proteins or tolipid molecules. Fructooligosaccharides are oligosaccharides consistingof short chains of fructose molecules.

Food sources of dietary fiber include, but are not limited to, grains,legumes, fruits, and vegetables. Grains providing dietary fiber include,but are not limited to, oats, rye, barley, wheat. Legumes providingfiber include, but are not limited to, peas and beans such as soybeans.Fruits and vegetables providing a source of fiber include, but are notlimited to, apples, oranges, pears, bananas, berries, tomatoes, greenbeans, broccoli, cauliflower, carrots, potatoes, celery. Plant foodssuch as bran, nuts, and seeds (such as flax seeds) are also sources ofdietary fiber. Parts of plants providing dietary fiber include, but arenot limited to, the stems, roots, leaves, seeds, pulp, and skin.

Although dietary fiber generally is derived from plant sources,indigestible animal products such as chitins are also classified asdietary fiber. Chitin is a polysaccharide composed of units ofacetylglucosamine joined by β(1-4) linkages, similar to the linkages ofcellulose.

Sources of dietary fiber often are divided into categories of solubleand insoluble fiber based on their solubility in water. Both soluble andinsoluble fibers are found in plant foods to varying degrees dependingupon the characteristics of the plant. Although insoluble in water,insoluble fiber has passive hydrophilic properties that help increasebulk, soften stools, and shorten transit time of fecal solids throughthe intestinal tract.

Unlike insoluble fiber, soluble fiber readily dissolves in water.Soluble fiber undergoes active metabolic processing via fermentation inthe colon, increasing the colonic microflora and thereby increasing themass of fecal solids. Fermentation of fibers by colonic bacteria alsoyields end-products with significant health benefits. For example,fermentation of the food masses produces gases and short-chain fattyacids. Acids produced during fermentation include butyric, acetic,propionic, and valeric acids that have various beneficial propertiessuch as stabilizing blood glucose levels by acting on pancreatic insulinrelease and providing liver control by glycogen breakdown. In addition,fiber fermentation may reduce atherosclerosis by lowering cholesterolsynthesis by the liver and reducing blood levels of LDL andtriglycerides. The acids produced during fermentation lower colonic pH,thereby protecting the colon lining from cancer polyp formation. Thelower colonic pH also increases mineral absorption, improves the barrierproperties of the colonic mucosal layer, and inhibits inflammatory andadhesion irritants. Fermentation of fibers also may benefit the immunesystem by stimulating production of T-helper cells, antibodies,leukocytes, splenocytes, cytokinins and lymphocytes.

Fatty Acid

In certain embodiments, the functional ingredient is at least one fattyacid. In one embodiment, a sweetener composition comprises at least onefatty acid, Reb X, and optionally at least one additive. In anotherembodiment, a sweetened composition comprises a sweetenable composition,at least one fatty acid, Reb X, and optionally at least one additive. Instill another embodiment, a sweetened composition comprises asweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one fatty acid, Reb X, andoptionally at least one additive.

As used herein, the at least one fatty acid may be single fatty acid ora plurality of fatty acids as a functional ingredient for the sweetenercomposition or sweetened compositions provided herein. Generally,according to particular embodiments of this invention, the at least onefatty acid is present in the sweetener composition or sweetenedcomposition in an amount sufficient to promote health and wellness.

As used herein, “fatty acid” refers to any straight chain monocarboxylicacid and includes saturated fatty acids, unsaturated fatty acids, longchain fatty acids, medium chain fatty acids, short chain fatty acids,fatty acid precursors (including omega-9 fatty acid precursors), andesterified fatty acids. As used herein, “long chain polyunsaturatedfatty acid” refers to any polyunsaturated carboxylic acid or organicacid with a long aliphatic tail. As used herein, “omega-3 fatty acid”refers to any polyunsaturated fatty acid having a first double bond asthe third carbon-carbon bond from the terminal methyl end of its carbonchain. In particular embodiments, the omega-3 fatty acid may comprise along chain omega-3 fatty acid. As used herein, “omega-6 fatty acid” anypolyunsaturated fatty acid having a first double bond as the sixthcarbon-carbon bond from the terminal methyl end of its carbon chain.

Suitable omega-3 fatty acids for use in embodiments of the presentinvention can be derived from algae, fish, animals, plants, orcombinations thereof, for example. Examples of suitable omega-3 fattyacids include, but are not limited to, linolenic acid, alpha-linolenicacid, eicosapentaenoic acid, docosahexaenoic acid, stearidonic acid,eicosatetraenoic acid and combinations thereof. In some embodiments,suitable omega-3 fatty acids can be provided in fish oils, (e.g.,menhaden oil, tuna oil, salmon oil, bonito oil, and cod oil), microalgaeomega-3 oils or combinations thereof. In particular embodiments,suitable omega-3 fatty acids may be derived from commercially availableomega-3 fatty acid oils such as Microalgae DHA oil (from Martek,Columbia, Md.), OmegaPure (from Omega Protein, Houston, Tex.), MarinolC-38 (from Lipid Nutrition, Channahon, Ill.), Bonito oil and MEG-3 (fromOcean Nutrition, Dartmouth, NS), Evogel (from Symrise, Holzminden,Germany), Marine Oil, from tuna or salmon (from Arista Wilton, Conn.),OmegaSource 2000, Marine Oil, from menhaden and Marine Oil, from cod(from OmegaSource, RTP, NC).

Suitable omega-6 fatty acids include, but are not limited to, linoleicacid, gamma-linolenic acid, dihommo-gamma-linolenic acid, arachidonicacid, eicosadienoic acid, docosadienoic acid, adrenic acid,docosapentacnoic acid and combinations thereof.

Suitable esterified fatty acids for embodiments of the present inventionmay include, but are not limited to, monoacylgycerols containing omega-3and/or omega-6 fatty acids, diacylgycerols containing omega-3 and/oromega-6 fatty acids, or triacylgycerols containing omega-3 and/oromega-6 fatty acids and combinations thereof.

Vitamin

In certain embodiments, the functional ingredient is at least onevitamin. In one embodiment, a sweetener composition comprises at leastone vitamin, Reb X, and optionally at least one additive. In anotherembodiment, a sweetened composition comprises a sweetenable composition,at least one vitamin, Reb X, and optionally at least one additive. Instill another embodiment, a sweetened composition comprises asweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one vitamin, Reb X, andoptionally at least one additive.

As used herein, the at least one vitamin may be single vitamin or aplurality of vitamins as a functional ingredient for the sweetener andsweetened compositions provided herein. Generally, according toparticular embodiments of this invention, the at least one vitamin ispresent in the sweetener composition or sweetened composition in anamount sufficient to promote health and wellness.

Vitamins are organic compounds that the human body needs in smallquantities for normal functioning. The body uses vitamins withoutbreaking them down, unlike other nutrients such as carbohydrates andproteins. To date, thirteen vitamins have been recognized, and one ormore can be used in the functional sweetener and sweetened compositionsherein. Suitable vitamins include, vitamin A, vitamin D, vitamin E,vitamin K, vitamin B1, vitamin B2, vitamin B3, vitamin B5, vitamin B6,vitamin B7, vitamin B9, vitamin B12, and vitamin C. Many of vitaminsalso have alternative chemical names, non-limiting examples of which areprovided below.

Vitamin Alternative names Vitamin A Retinol Retinaldehyde Retinoic acidRetinoids Retinal Retinoic ester Vitamin D (vitamins Calciferol D1-D5)Cholecalciferol Lumisterol Ergocalciferol Dihydrotachysterol7-dehydrocholesterol Vitamin E Tocopherol Tocotrienol Vitamin KPhylloquinone Naphthoquinone Vitamin B1 Thiamin Vitamin B2 RiboflavinVitamin G Vitamin B3 Niacin Nicotinic acid Vitamin PP Vitamin B5Pantothenic acid Vitamin B6 Pyridoxine Pyridoxal Pyridoxamine Vitamin B7Biotin Vitamin H Vitamin B9 Folic acid Folate Folacin Vitamin MPteroyl-L-glutamic acid Vitamin B12 Cobalamin Cyanocobalamin Vitamin CAscorbic acid

Various other compounds have been classified as vitamins by someauthorities. These compounds may be termed pseudo-vitamins and include,but are not limited to, compounds such as ubiquinone (coenzyme Q10),pangamic acid, dimethylglycine, taestrile, amygdaline, flavanoids,para-aminobenzoic acid, adenine, adenylic acid, and s-methylmethionine.As used herein, the term vitamin includes pseudo-vitamins.

In some embodiments, the vitamin is a fat-soluble vitamin chosen fromvitamin A, D, E, K and combinations thereof.

In other embodiments, the vitamin is a water-soluble vitamin chosen fromvitamin B1, vitamin B2, vitamin B3, vitamin B6, vitamin B12, folic acid,biotin, pantothenic acid, vitamin C and combinations thereof.

Glucosamine

In certain embodiments, the functional ingredient is glucosamine. In oneembodiment, a sweetener composition comprises glucosamine, Reb X, andoptionally at least one additive. In another embodiment, a sweetenedcomposition comprises a sweetenable composition, glucosamine, Reb X, andoptionally at least one additive. In still another embodiment, asweetened composition comprises a sweetenable composition and asweetener composition, wherein the sweetener composition comprisesglucosamine, Reb X, and optionally at least one additive.

Generally, according to particular embodiments of this invention,glucosamine is present in the functional sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

Glucosamine, also called chitosamine, is an amino sugar that is believedto be an important precursor in the biochemical synthesis ofglycosylated proteins and lipids. D-glucosamine occurs naturally in thecartilage in the form of glucosamine-6-phosphate, which is synthesizedfrom fructose-6-phosphate and glutamine. However, glucosamine also isavailable in other forms, non-limiting examples of which includeglucosamine hydrochloride, glucosamine sulfate, N-acetyl-glucosamine, orany other salt forms or combinations thereof. Glucosamine may beobtained by acid hydrolysis of the shells of lobsters, crabs, shrimps,or prawns using methods well known to those of ordinary skill in theart. In a particular embodiment, glucosamine may be derived from fungalbiomass containing chitin, as described in U.S. Patent Publication No.2006/0172392.

The sweetener compositions or sweetened composition can further comprisechondroitin sulfate.

Mineral

In certain embodiments, the functional ingredient is at least onemineral. In one embodiment, a sweetener composition comprises at leastone mineral, Reb X, and optionally at least one additive. In anotherembodiment, a sweetened composition comprises a sweetenable composition,at least one mineral, Reb X, and optionally at least one additive. Instill another embodiment, a sweetened composition comprises asweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one mineral, Reb X, andoptionally at least one additive.

As used herein, the at least one mineral may be single mineral or aplurality of minerals as a functional ingredient for the sweetenercompositions or sweetened compositions provided herein. Generally,according to particular embodiments of this invention, the at least onemineral is present in the sweetener composition or sweetened compositionin an amount sufficient to promote health and wellness.

Minerals, in accordance with the teachings of this invention, compriseinorganic chemical elements required by living organisms. Minerals arecomprised of a broad range of compositions (e.g., elements, simplesalts, and complex silicates) and also vary broadly in crystallinestructure. They may naturally occur in foods and beverages, may be addedas a supplement, or may be consumed or administered separately fromfoods or beverages.

Minerals may be categorized as either bulk minerals, which are requiredin relatively large amounts, or trace minerals, which are required inrelatively small amounts. Bulk minerals generally are required inamounts greater than or equal to about 100 mg per day and trace mineralsare those that are required in amounts less than about 100 mg per day.

In particular embodiments of this invention, the mineral is chosen frombulk minerals, trace minerals or combinations thereof. Non-limitingexamples of bulk minerals include calcium, chlorine, magnesium,phosphorous, potassium, sodium, and sulfur. Non-limiting examples oftrace minerals include chromium, cobalt, copper, fluorine, iron,manganese, molybdenum, selenium, zinc, and iodine. Although iodinegenerally is classified as a trace mineral, it is required in largerquantities than other trace minerals and often is categorized as a bulkmineral.

In other particular embodiments of this invention, the mineral is atrace mineral, believed to be necessary for human nutrition,non-limiting examples of which include bismuth, boron, lithium, nickel,rubidium, silicon, strontium, tellurium, tin, titanium, tungsten, andvanadium.

The minerals embodied herein may be in any form known to those ofordinary skill in the art. For example, in a particular embodiment theminerals may be in their ionic form, having either a positive ornegative charge. In another particular embodiment the minerals may be intheir molecular form. For example, sulfur and phosphorous often arefound naturally as sulfates, sulfides, and phosphates.

Preservative

In certain embodiments, the functional ingredient is at least onepreservative. In one embodiment, a sweetener composition comprises atleast one preservative, Reb X, and optionally at least one additive. Inanother embodiment, a sweetened composition comprises a sweetenablecomposition, at least one preservative, Reb X, and optionally at leastone additive. In still another embodiment, a sweetened compositioncomprises a sweetenable composition and a sweetener composition, whereinthe sweetener composition comprises at least one preservative, Reb X,and optionally at least one additive.

As used herein, the at least one preservative may be single preservativeor a plurality of preservatives as a functional ingredient for thesweetener compositions or sweetened composition provided herein.Generally, according to particular embodiments of this invention, the atleast one preservative is present in the sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

In particular embodiments of this invention, the preservative is chosenfrom antimicrobials, antioxidants, antienzymatics or combinationsthereof. Non-limiting examples of antimicrobials include sulfites,propionates, benzoates, sorbates, nitrates, nitrites, bacteriocins,salts, sugars, acetic acid, dimethyl dicarbonate (DMDC), ethanol, andozone.

According to a particular embodiment, the preservative is a sulfite.Sulfites include, but are not limited to, sulfur dioxide, sodiumbisulfite, and potassium hydrogen sulfite.

According to another particular embodiment, the preservative is apropionate. Propionates include, but are not limited to, propionic acid,calcium propionate, and sodium propionate.

According to yet another particular embodiment, the preservative is abenzoate. Benzoates include, but are not limited to, sodium benzoate andbenzoic acid.

In another particular embodiment, the preservative is a sorbate.Sorbates include, but are not limited to, potassium sorbate, sodiumsorbate, calcium sorbate, and sorbic acid.

In still another particular embodiment, the preservative is a nitrateand/or a nitrite. Nitrates and nitrites include, but are not limited to,sodium nitrate and sodium nitrite.

In yet another particular embodiment, the at least one preservative is abacteriocin, such as, for example, raisin.

In another particular embodiment, the preservative is ethanol.

In still another particular embodiment, the preservative is ozone.

Non-limiting examples of antienzymatics suitable for use aspreservatives in particular embodiments of the invention includeascorbic acid, citric acid, and metal chelating agents such asethylenediaminetetraacetic acid (EDTA).

Hydration Agent

In certain embodiments, the functional ingredient is at least onehydration agent. In one embodiment, a sweetener composition comprises atleast one hydration agent, Reb X, and optionally at least one additive.In another embodiment, a sweetened composition comprises a sweetenablecomposition, at least one hydration agent, Reb X, and optionally atleast one additive. In still another embodiment, a sweetened compositioncomprises a sweetenable composition and a sweetener composition, whereinthe sweetener composition comprises at least one hydration gent, Reb X,and optionally at least one additive.

As used herein, the at least one hydration agent may be single hydrationagent or a plurality of hydration agents as a functional ingredient forthe sweetener compositions or sweetened composition provided herein.Generally, according to particular embodiments of this invention, the atleast one hydration agent is present in the sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

Hydration products help the body to replace fluids that are lost throughexcretion. For example, fluid is lost as sweat in order to regulate bodytemperature, as urine in order to excrete waste substances, and as watervapor in order to exchange gases in the lungs. Fluid loss can also occurdue to a wide range of external causes, non-limiting examples of whichinclude physical activity, exposure to dry air, diarrhea, vomiting,hyperthermia, shock, blood loss, and hypotension. Diseases causing fluidloss include diabetes, cholera, gastroenteritis, shigellosis, and yellowfever. Forms of malnutrition that cause fluid loss include the excessiveconsumption of alcohol, electrolyte imbalance, fasting, and rapid weightloss.

In a particular embodiment, the hydration product is a composition thathelps the body replace fluids that are lost during exercise.Accordingly, in a particular embodiment, the hydration product is anelectrolyte, non-limiting examples of which include sodium, potassium,calcium, magnesium, chloride, phosphate, bicarbonate, and combinationsthereof. Suitable electrolytes for use in particular embodiments of thisinvention are also described in U.S. Pat. No. 5,681,569, the disclosureof which is expressly incorporated herein by reference. In particularembodiments, the electrolytes are obtained from their correspondingwater-soluble salts. Non-limiting examples of salts for use inparticular embodiments include chlorides, carbonates, sulfates,acetates, bicarbonates, citrates, phosphates, hydrogen phosphates,tartates, sorbates, citrates, benzoates, or combinations thereof. Inother embodiments, the electrolytes are provided by juice, fruitextracts, vegetable extracts, tea, or teas extracts.

In particular embodiments of this invention, the hydration product is acarbohydrate to supplement energy stores burned by muscles. Suitablecarbohydrates for use in particular embodiments of this invention aredescribed in U.S. Pat. Nos. 4,312,856, 4,853,237, 5,681,569, and6,989,171, the disclosures of which are expressly incorporated herein byreference. Non-limiting examples of suitable carbohydrates includemonosaccharides, disaccharides, oligosaccharides, complexpolysaccharides or combinations thereof. Non-limiting examples ofsuitable types of monosaccharides for use in particular embodimentsinclude trioses, tetroses, pentoses, hexoses, heptoses, octoses, andnonoses. Non-limiting examples of specific types of suitablemonosaccharides include glyceraldehyde, dihydroxyacetone, erythrose,threose, erythrulose, arabinose, lyxose, ribose, xylose, ribulose,xylulose, allose, altrose, galactose, glucose, gulose, idose, mannose,talose, fructose, psicose, sorbose, tagatose, mannoheptulose,sedoheltulose, octolose, and sialose. Non-limiting examples of suitabledisaccharides include sucrose, lactose, and maltose. Non-limitingexamples of suitable oligosaccharides include saccharose, maltotriose,and maltodextrin. In other particular embodiments, the carbohydrates areprovided by a corn syrup, a beet sugar, a cane sugar, a juice, or a tea.

In another particular embodiment, the hydration is a flavanol thatprovides cellular rehydration. Flavanols are a class of naturalsubstances present in plants, and generally comprise a2-phenylbenzopyrone molecular skeleton attached to one or more chemicalmoieties. Non-limiting examples of suitable flavanols for use inparticular embodiments of this invention include catechin, epicatechin,gallocatechin, epigallocatechin, epicatechin gallate, epigallocatechin3-gallate, theaflavin, theaflavin 3-gallate, theaflavin 3′-gallate,theaflavin 3,3′ gallate, thearubigin or combinations thereof. Severalcommon sources of flavanols include tea plants, fruits, vegetables, andflowers. In preferred embodiments, the flavanol is extracted from greentea.

In a particular embodiment, the hydration product is a glycerol solutionto enhance exercise endurance. The ingestion of a glycerol containingsolution has been shown to provide beneficial physiological effects,such as expanded blood volume, lower heart rate, and lower rectaltemperature.

Probiotics/Prebiotics

In certain embodiments, the functional ingredient is chosen from atleast one probiotic, prebiotic and combination thereof. In oneembodiment, a sweetener composition comprises at least one probiotic,prebiotic and combination thereof; Reb X; and optionally at least oneadditive. In another embodiment, a sweetened composition comprises asweetenable composition, at least one at least one probiotic, prebioticand combination thereof; Reb X; and optionally at least one additive. Instill another embodiment, a sweetened composition comprises asweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one probiotic, prebiotic andcombination thereof; Reb X; and optionally at least one additive.

As used herein, the at least one probiotic or prebiotic may be singleprobiotic or prebiotic or a plurality of probiotics or prebiotics as afunctional ingredient for the sweetener compositions or sweetenedcomposition provided herein. Generally, according to particularembodiments of this invention, the at least one probiotic, prebiotic orcombination thereof is present in the sweetener composition or sweetenedcomposition in an amount sufficient to promote health and wellness.

Probiotics, in accordance with the teachings of this invention, comprisemicroorganisms that benefit health when consumed in an effective amount.Desirably, probiotics beneficially affect the human body'snaturally-occurring gastrointestinal microflora and impart healthbenefits apart from nutrition. Probiotics may include, withoutlimitation, bacteria, yeasts, and fungi.

According to particular embodiments, the probiotic is a beneficialmicroorganisms that beneficially affects the human body'snaturally-occurring gastrointestinal microflora and imparts healthbenefits apart from nutrition. Examples of probiotics include, but arenot limited to, bacteria of the genus Lactobacilli, Bifidobacteria,Streptococci, or combinations thereof, that confer beneficial effects tohumans.

In particular embodiments of the invention, the at least one probioticis chosen from the genus Lactobacilli. Lactobacilli (i.e., bacteria ofthe genus Lactobacillus, hereinafter “L.”) have been used for severalhundred years as a food preservative and for promoting human health.Non-limiting examples of species of Lactobacilli found in the humanintestinal tract include L. acidophilus, L. casei, L. fermentum, L.saliva roes, L. brevis, L. leichmannii, L. plantarum, L. cellobiosus, L.reuteri, L. rhamnosus, L. GG, L. bulgaricus, and L. thermophilus.

According to other particular embodiments of this invention, theprobiotic is chosen from the genus Bifidobacteria. Bifidobacteria alsoare known to exert a beneficial influence on human health by producingshort chain fatty acids (e.g., acetic, propionic, and butyric acids),lactic, and formic acids as a result of carbohydrate metabolism.Non-limiting species of Bifidobacteria found in the humangastrointestinal tract include B. angulatum, B. animalis, B. asteroides,B. bifidum, B. bourn, B. breve, B. catenulatum, B. choerinum, B.coryneforme, B. cuniculi, B. dentium, B. gallicum, B. gallinarum, Bindicum, B. longum, B. magnum, B. merycicum, B. minimum, B.pseudocatenulatum, B. pseudolongum, B. psychraerophilum, B. pullorum, B.ruminantium, B. saeculare, B. scardovii, B. simiae, B. subtile, B.thermacidophilum, B. thermophilum, B. urinalis, and B. sp.

According to other particular embodiments of this invention, theprobiotic is chosen from the genus Streptococcus. Streptococcusthermophilus is a gram-positive facultative anaerobe. It is classifiedas a lactic acid bacteria and commonly is found in milk and milkproducts, and is used in the production of yogurt. Other non-limitingprobiotic species of this bacteria include Streptococcus salivarus andStreptococcus cremoris.

Probiotics that may be used in accordance with this invention arewell-known to those of skill in the art. Non-limiting examples offoodstuffs comprising probiotics include yogurt, sauerkraut, ketir,kimchi, fermented vegetables, and other foodstuffs containing amicrobial element that beneficially affects the host animal by improvingthe intestinal microbalance.

Prebiotics, in accordance with the teachings of this invention, arecompositions that promote the growth of beneficial bacteria in theintestines. Prebiotic substances can be consumed by a relevantprobiotic, or otherwise assist in keeping the relevant probiotic aliveor stimulate its growth. When consumed in an effective amount,prebiotics also beneficially affect the human body's naturally-occurringgastrointestinal microflora and thereby impart health benefits apartfrom just nutrition. Prebiotic foods enter the colon and serve assubstrate for the endogenous bacteria, thereby indirectly providing thehost with energy, metabolic substrates, and essential micronutrients.The body's digestion and absorption of prebiotic foods is dependent uponbacterial metabolic activity, which salvages energy for the host fromnutrients that escaped digestion and absorption in the small intestine.

Prebiotics, in accordance with the embodiments of this invention,include, without limitation, mucopolysaccharides, oligosaccharides,polysaccharides, amino acids, vitamins, nutrient precursors, proteinsand combinations thereof.

According to a particular embodiment of this invention, the prebiotic ischosen from dietary fibers, including, without limitation,polysaccharides and oligosaccharides. These compounds have the abilityto increase the number of probiotics, which leads to the benefitsconferred by the probiotics. Non-limiting examples of oligosaccharidesthat are categorized as prebiotics in accordance with particularembodiments of this invention include fructooligosaceharides, insulins,isomalto-oligosaccharides, lactilol, lactosucrose, lactulose,pyrodextrins, soy oligosaccharides, transgalacto-oligosaccharides, andxylo-oligosaccharides.

According to other particular embodiments of the invention, theprebiotic is an amino acid. Although a number of known prebiotics breakdown to provide carbohydrates for probiotics, some probiotics alsorequire amino acids for nourishment.

Prebiotics are found naturally in a variety of foods including, withoutlimitation, bananas, berries, asparagus, garlic, wheat, oats, barley(and other whole grains), flaxseed, tomatoes, Jerusalem artichoke,onions and chicory, greens (e.g., dandelion greens, spinach, collardgreens, chard, kale, mustard greens, turnip greens), and legumes (e.g.,lentils, kidney beans, chickpeas, navy beans, white beans, black beans).

Weight Management Agent

In certain embodiments, the functional ingredient is at least one weightmanagement agent. In one embodiment, a sweetener composition comprisesat least one weight management agent, Reb X, and optionally at least oneadditive. In another embodiment, a sweetened composition comprises asweetenable composition, at least one weight management agent, Reb X,and optionally at least one additive. In still another embodiment, asweetened composition comprises a sweetenable composition and asweetener composition, wherein the sweetener composition comprises atleast one weight management agent, Reb X, and optionally at least oneadditive.

As used herein, the at least one weight management agent may be singleweight management agent or a plurality of weight management agents as afunctional ingredient for the sweetener compositions or sweetenedcomposition provided herein. Generally, according to particularembodiments of this invention, the at least one weight management agentis present in the sweetener composition or sweetened composition in anamount sufficient to promote health and wellness.

As used herein, “a weight management agent” includes an appetitesuppressant and/or a thermogenesis agent. As used herein, the phrases“appetite suppressant”, “appetite satiation compositions”, “satietyagents”, and “satiety ingredients” are synonymous. The phrase “appetitesuppressant” describes macronutrients, herbal extracts, exogenoushormones, anorectics, anorexigenics, pharmaceutical drugs, andcombinations thereof, that when delivered in an effective amount,suppress, inhibit, reduce, or otherwise curtail a person's appetite. Thephrase “thermogenesis agent” describes macronutrients, herbal extracts,exogenous hormones, anorectics, anorexigenics, pharmaceutical drugs, andcombinations thereof, that when delivered in an effective amount,activate or otherwise enhance a person's thermogenesis or metabolism.

Suitable weight management agents include macronutrient selected fromthe group consisting of proteins, carbohydrates, dietary fats, andcombinations thereof. Consumption of proteins, carbohydrates, anddietary fats stimulates the release of peptides withappetite-suppressing effects. For example, consumption of proteins anddietary fats stimulates the release of the gut hormone cholecytokinin(CCK), while consumption of carbohydrates and dietary fats stimulatesrelease of Glucagon-like peptide 1 (GLP-1).

Suitable macronutrient weight management agents also includecarbohydrates. Carbohydrates generally comprise sugars, starches,cellulose and gums that the body converts into glucose for energy.Carbohydrates often are classified into two categories, digestiblecarbohydrates (e.g., monosaccharides, disaccharides, and starch) andnon-digestible carbohydrates (e.g., dietary fiber). Studies have shownthat non-digestible carbohydrates and complex polymeric carbohydrateshaving reduced absorption and digestibility in the small intestinestimulate physiologic responses that inhibit food intake. Accordingly,the carbohydrates embodied herein desirably comprise non-digestiblecarbohydrates or carbohydrates with reduced digestibility. Non-limitingexamples of such carbohydrates include polydextrose; inulin;monosaccharide-derived polyols such as erythritol, mannitol, xylitol,and sorbitol; disaccharide-derived alcohols such as isomalt, lactitol,and maltitol; and hydrogenated starch hydrolysates. Carbohydrates aredescribed in more detail herein below.

In another particular embodiment weight management agent is a dietaryfat. Dietary fats are lipids comprising combinations of saturated andunsaturated fatty acids. Polyunsaturated fatty acids have been shown tohave a greater satiating power than mono-unsaturated fatty acids.Accordingly, the dietary fats embodied herein desirably comprisepoly-unsaturated fatty acids, non-limiting examples of which includetriacylglycerols.

In a particular embodiment, the weight management agents is an herbalextract. Extracts from numerous types of plants have been identified aspossessing appetite suppressant properties. Non-limiting examples ofplants whose extracts have appetite suppressant properties includeplants of the genus Hoodia, Trichocaulon, Caralluma, Stapelia, Orbea,Asclepias, and Camelia. Other embodiments include extracts derived fromGymnema Sylvestre, Kola Nut, Citrus Auran tium, Yerba Mate, GriffoniaSimplicifolia, Guarana, myrrh, guggul Lipid, and black current seed oil.

The herbal extracts may be prepared from any type of plant material orplant biomass. Non-limiting examples of plant material and biomassinclude the stems, roots, leaves, dried powder obtained from the plantmaterial, and sap or dried sap. The herbal extracts generally areprepared by extracting sap from the plant and then spray-drying the sap.Alternatively, solvent extraction procedures may be employed. Followingthe initial extraction, it may be desirable to further fractionate theinitial extract (e.g., by column chromatography) in order to obtain anherbal extract with enhanced activity. Such techniques are well known tothose of ordinary skill in the art.

In a particular embodiment, the herbal extract is derived from a plantof the genus Hoodia, species of which include H. alstonii, H. currorii,H. dregei, H. flava, H. gordonii, H. jutatae, H. mossamedensis, H.officinalis, H. parviflorai, H. pedicellata, H. pilifera, H. ruschii,and H. triebneri. Hoodia plants are stem succulents native to southernAfrica. A sterol glycoside of Hoodia, known as P57, is believed to beresponsible for the appetite-suppressant effect of the Hoodia species.

In another particular embodiment, the herbal extract is derived from aplant of the genus Caralluma, species of which include C. indica, C.fimbriata, C. attenuate, C. tuberculate, C. edulis, C. adscendens, C.stalagmifera, C. umbellate, C. penicillata, C. russeliana, C.retrospicens, C. Arabica, and C. lasiantha. Carralluma plants belong tothe same Subfamily as Hoodia, Asclepiadaceae. Caralluma are small, erectand fleshy plants native to India having medicinal properties, such asappetite suppression, that generally are attributed to glycosidesbelonging to the pregnane group of glycosides, non-limiting examples ofwhich include caratuberside A, caratuberside B, bouceroside I,bouceroside II, bouceroside III, bouceroside IV, bouceroside V,bouceroside VI, bouceroside VII, bouceroside VIII, bouceroside IX, andbouceroside X.

In another particular embodiment, the at least one herbal extract isderived from a plant of the genus Trichocaulon. Trichocaulon plants aresucculents that generally are native to southern Africa, similar toHoodia, and include the species T. piliferum and T. officinale.

In another particular embodiment, the herbal extract is derived from aplant of the genus Stapelia or Orbea, species of which include S.gigantean and O. variegate, respectively. Both Stapelia and Orbea plantsbelong to the same Subfamily as Hoodia, Asclepiadaceae. Not wishing tobe bound by any theory, it is believed that the compounds exhibitingappetite suppressant activity are saponins, such as pregnane glycosides,which include stavarosides A, B, C, D, E, F, G, H, I, J, and K.

In another particular embodiment, the herbal extract is derived from aplant of the genus Asclepias. Asclepias plants also belong to theAsclepiadaceae family of plants. Non-limiting examples of Asclepiasplants include A. incarnate, A. curassayica, A. syriaca, and A.tuberose. Not wishing to be bound by any theory, it is believed that theextracts comprise steroidal compounds, such as pregnane glycosides andpregnane aglycone, having appetite suppressant effects.

In a particular embodiment, the weight management agent is an exogenoushormone having a weight management effect. Non-limiting examples of suchhormones include CCK, peptide YY, ghrelin, bombesin andgastrin-releasing peptide (GRP), enterostatin, apolipoprotein A-IV,CLP-1, amylin, somastatin, and leptin.

In another embodiment, the weight management agent is a pharmaceuticaldrug. Non-limiting examples include phentenime, diethylpropion,phendimetrazine, sibutramine, rimonabant, oxyntomodulin, floxetinehydrochloride, ephedrine, phenethylamine, or other stimulants.

The at least one weight management agent may be utilized individually orin combination as a functional ingredient for the sweetener compositionsprovided in this invention.

Osteoporosis Management Agent

In certain embodiments, the functional ingredient is at least oneosteoporosis management agent. In one embodiment, a sweetenercomposition comprises at least one osteoporosis management agent, Reb X,and optionally at least one additive. In another embodiment, a sweetenedcomposition comprises a sweetenable composition, at least oneosteoporosis management agent, Reb X, and optionally at least oneadditive. In still another embodiment, a sweetened composition comprisesa sweetenable composition and a sweetener composition, wherein thesweetener composition comprises at least one osteoporosis managementagent, Reb X, and optionally at least one additive.

As used herein, the at least one osteoporosis management agent may besingle osteoporosis management agent or a plurality of osteoporosismanagement agent as a functional ingredient for the sweetenercompositions or sweetened composition provided herein. Generally,according to particular embodiments of this invention, the at least oneosteoporosis management agent is present in the sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

Osteoporosis is a skeletal disorder of compromised bone strength,resulting in an increased risk of bone fracture. Generally, osteoporosisis characterized by reduction of the bone mineral density (BMD),disruption of bone micro-architecture, and changes to the amount andvariety of non-collagenous proteins in the bone.

In certain embodiments, the osteoporosis management agent is at leastone calcium source. According to a particular embodiment, the calciumsource is any compound containing calcium, including salt complexes,solubilized species, and other forms of calcium. Non-limiting examplesof calcium sources include amino acid chelated calcium, calciumcarbonate, calcium oxide, calcium hydroxide, calcium sulfate, calciumchloride, calcium phosphate, calcium hydrogen phosphate, calciumdihydrogen phosphate, calcium citrate, calcium malate, calcium citratemalate, calcium gluconate, calcium tartrate, calcium lactate,solubilized species thereof, and combinations thereof.

According to a particular embodiment, the osteoporosis management agentis a magnesium source. The magnesium source is any compound containingmagnesium, including salt complexes, solubilized species, and otherforms of magnesium. Non-limiting examples of magnesium sources includemagnesium chloride, magnesium citrate, magnesium gluceptate, magnesiumgluconate, magnesium lactate, magnesium hydroxide, magnesium picolate,magnesium sulfate, solubilized species thereof, and mixtures thereof. Inanother particular embodiment, the magnesium source comprises an aminoacid chelated or creatine chelated magnesium.

In other embodiments, the osteoporosis agent is chosen from vitamins D,C, K, their precursors and/or beta-carotene and combinations thereof

Numerous plants and plant extracts also have been identified as beingeffective in the prevention and treatment of osteoporosis. Not wishingto be bound by any theory, it is believed that the plants and plantextracts stimulates bone morphogenic proteins and/or inhibits boneresorption, thereby stimulating bone regeneration and strength.Non-limiting examples of suitable plants and plant extracts asosteoporosis management agents include species of the genus Taraxacumand Amelanchier, as disclosed in U.S. Patent Publication No.2005/0106215, and species of the genus Lindera, Artemisia, Acorus,Carthamus, Carum, Cnidium, Curcuma, Cyperus, Juniperus, Prunus, Iris,Cichorium, Dodonaea, Epimedium, Erigonoum, Soya, Mentha, Ocimum, thymus,Tanacetum, Plantago, Spearmint, Bixa, Vitis, Rosemarinus, Rhus, andAnethum, as disclosed in U.S. Patent Publication No. 2005/0079232.

Phytoestrogen

In certain embodiments, the functional ingredient is at least onephytoestrogen. In one embodiment, a sweetener composition comprises atleast one phytoestrogen, Reb X, and optionally at least one additive. Inanother embodiment, a sweetened composition comprises a sweetenablecomposition, at least one phytoestrogen, Reb X, and optionally at leastone additive. In still another embodiment, a sweetened compositioncomprises a sweetenable composition and a sweetener composition, whereinthe sweetener composition comprises at least one phytoestrogen, Reb X,and optionally at least one additive.

As used herein, the at least one phytoestrogen may be singlephytoestrogen or a plurality of phytoestrogens as a functionalingredient for the sweetener compositions or sweetened compositionprovided herein. Generally, according to particular embodiments of thisinvention, the at least one phytoestrogen is present in the sweetenercomposition or sweetened composition in an amount sufficient to promotehealth and wellness.

Phytoestrogens are compounds found in plants which can typically bedelivered into human bodies by ingestion of the plants or the plantparts having the phytoestrogens. As used herein, “phytoestrogen” refersto any substance which, when introduced into a body causes anestrogen-like effect of any degree. For example, a phytoestrogen maybind to estrogen receptors within the body and have a smallestrogen-like effect.

Examples of suitable phytoestrogens for embodiments of this inventioninclude, but are not limited to, isoflavones, stilbenes, lignans,resorcyclic acid lactones, coumestans, coumestrol, equol, andcombinations thereof. Sources of suitable phytoestrogens include, butare not limited to, whole grains, cereals, fibers, fruits, vegetables,black cohosh, agave root, black currant, black haw, chasteberries, crampbark, dong quai root, devil's club root, false unicorn root, ginsengroot, groundsel herb, licorice, liferoot herb, motherwort herb, peonyroot, raspberry leaves, rose family plants, sage leaves, sarsaparillaroot, saw palmetto berried, wild yam root, yarrow blossoms, legumes,soybeans, soy products (e.g., miso, soy flour, soymilk, soy nuts, soyprotein isolate, tempen, or tofu) chick peas, nuts, lentils, seeds,clover, red clover, dandelion leaves, dandelion roots, fenugreek seeds,green tea, hops, red wine, flaxseed, garlic, onions, linseed, borage,butterfly weed, caraway, chaste tree, vitex, dates, dill, fennel seed,gotu kola, milk thistle, pennyroyal, pomegranates, southernwood, soyaflour, tansy, and root of the kudzu vine (pueraria root) and the like,and combinations thereof.

Isoflavones belong to the group of phytonutrients called polyphenols. Ingeneral, polyphenols (also known as “polyphenolics”), are a group ofchemical substances found in plants, characterized by the presence ofmore than one phenol group per molecule.

Suitable phytoestrogen isoflavones in accordance with embodiments ofthis invention include genistein, daidzein, glycitein, biochanin A,formononetin, their respective naturally occurring glycosides andglycoside conjugates, matairesinol, secoisolariciresinol, enterolactone,enterodiol, textured vegetable protein, and combinations thereof.

Suitable sources of isoflavones for embodiments of this inventioninclude, but are not limited to, soy beans, soy products, legumes,alfalfa spouts, chickpeas, peanuts, and red clover.

Long-Chain Primary Aliphatic Saturated Alcohol

In certain embodiments, the functional ingredient is at least one longchain primary aliphatic saturated alcohol. In one embodiment, asweetener composition comprises at least one long chain primaryaliphatic saturated alcohol, Reb X, and optionally at least oneadditive. In another embodiment, a sweetened composition comprises asweetenable composition, at least one long chain primary aliphaticsaturated alcohol, Reb X, and optionally at least one additive. In stillanother embodiment, a sweetened composition comprises a sweetenablecomposition and a sweetener composition, wherein the sweetenercomposition comprises at least one long chain primary aliphaticsaturated alcohol, Reb X, and optionally at least one additive.

As used herein, the at least one long chain primary aliphatic saturatedalcohol may be single long chain primary aliphatic saturated alcohol ora plurality of long chain primary aliphatic saturated alcohols as afunctional ingredient for the sweetener compositions or sweetenedcomposition provided herein. Generally, according to particularembodiments of this invention, the at least one long chain primaryaliphatic saturated alcohol is present in the sweetener composition orsweetened composition in an amount sufficient to promote health andwellness.

Long-chain primary aliphatic saturated alcohols are a diverse group oforganic compounds. The term alcohol refers to the fact these compoundsfeature a hydroxyl group (—OH) bound to a carbon atom. The term primaryrefers to the fact that in these compounds the carbon atom which isbound to the hydroxyl group is bound to only one other carbon atom. Theterm saturated refers to the fact that these compounds feature no carbonto carbon pi bonds. The term aliphatic refers to the fact that thecarbon atoms in these compounds are joined together in straight orbranched chains rather than in rings. The term long-chain refers to thefact that the number of carbon atoms in these compounds is at least 8carbons).

Non-limiting examples of particular long-chain primary aliphaticsaturated alcohols for use in particular embodiments of the inventioninclude the 8 carbon atom 1-octanol, the 9 carbon 1-nonanol, the 10carbon atom 1-decanol, the 12 carbon atom 1-dodecanol, the 14 carbonatom 1-tetradecanol, the 16 carbon atom 1-hexadecanol, the 18 carbonatom 1-octadecanol, the 20 carbon atom 1-eicosanol, the 22 carbon1-docosanol, the 24 carbon 1-tetracosanol, the 26 carbon 1-hexacosanol,the 27 carbon 1-heptacosanol, the 28 carbon 1-octanosol, the 29 carbon1-nonacosanol, the 30 carbon 1-triacontanol, the 32 carbon1-dotriacontanol, and the 34 carbon 1-tetracontanol.

In a particularly desirable embodiment of the invention, the long-chainprimary aliphatic saturated alcohols are policosanol. Policosanol is theterm for a mixture of long-chain primary aliphatic saturated alcoholscomposed primarily of 28 carbon 1-octanosol and 30 carbon1-triacontanol, as well as other alcohols in lower concentrations suchas 22 carbon 1-docosanol, 24 carbon 1-tetracosanol, 26 carbon1-hexacosanol, 27 carbon 1-heptacosanol, 29 carbon 1-nonacosanol, 32carbon 1-dotriacontanol, and 34 carbon 1-tetracontanol.

Long-chain primary aliphatic saturated alcohols are derived from naturalfats and oils. They may be obtained from these sources by usingextraction techniques well known to those of ordinary skill in the art.Policosanols can be isolated from a variety of plants and materialsincluding sugar cane (Saccharum officinarium), yams (e.g. Dioscoreaopposite), bran from rice (e.g. Oryza sativa), and beeswax. Policosanolsmay be obtained from these sources by using extraction techniques wellknown to those of ordinary skill in the art. A description of suchextraction techniques can be found in U.S. Pat. Appl. No. 2005/0220868,the disclosure of which is expressly incorporated by reference.

Phytosterols

In certain embodiments, the functional ingredient is at least onephytosterol, phytostanol or combination thereof. In one embodiment, asweetener composition comprises at least one phytosterol, phytostanol orcombination thereof. Reb X; and optionally at least one additive. Inanother embodiment, a sweetened composition comprises a sweetenablecomposition, at least one phytosterol, phytostanol or combinationthereof; Reb X; and optionally, at least one additive. In still anotherembodiment, a sweetened composition comprises a sweetenable compositionand a sweetener composition, wherein the sweetener composition comprisesat least one phytosterol, phytostanol or combination thereof; Reb X; andoptionally at least one additive.

Generally, according to particular embodiments of this invention, the atleast one phytosterol, phytostanol or combination thereof is present inthe sweetener composition or sweetened composition in an amountsufficient to promote health and wellness.

As used herein, the phrases “stanol”, “plant stanol” and “phytostanol”are synonymous.

Plant sterols and stanols are present naturally in small quantities inmany fruits, vegetables, nuts, seeds, cereals, legumes, vegetable oils,bark of the trees and other plant sources. Although people normallyconsume plant sterols and stanols every day, the amounts consumed areinsufficient to have significant cholesterol-lowering effects or otherhealth benefits. Accordingly, it would be desirable to supplement foodand beverages with plant sterols and stanols.

Sterols are a subgroup of steroids with a hydroxyl group at C-3.Generally, phytosterols have a double bond within the steroid nucleus,like cholesterol; however, phytosterols also may comprise a substitutedsidechain (R) at C-24, such as an ethyl or methyl group, or anadditional double bond. The structures of phytosterols are well known tothose of skill in the art.

At least 44 naturally-occurring phytosterols have been discovered, andgenerally are derived from plants, such as corn, soy, wheat, and woodoils; however, they also may be produced synthetically to formcompositions identical to those in nature or having properties similarto those of naturally-occurring phytosterols. According to particularembodiments of this invention, non-limiting examples of phytosterolswell known to those or ordinary skill in the art include4-desmethylsterols (e.g., β-sitosterol, campesterol, stigmasterol,brassicasterol, 22-dehydrobrassicasterol, and Δ5-avenasterol),4-monomethyl sterols, and 4,4-dimethyl sterols (triterpene alcohols)(e.g., cycloartenol, 24-methylenecycloartanol, and cyclobranol).

As used herein, the phrases “stanol”, “plant stanol” and “phytostanol”are synonymous. Phytostanols are saturated sterol alcohols present inonly trace amounts in nature and also may be synthetically produced,such as by hydrogenation of phytosterols. According to particularembodiments of this invention, non-limiting examples of phytostanolsinclude β-sitostanol, campestanol, cycloartanol, and saturated forms ofother triterpene alcohols.

Both phytosterols and phytostanols, as used herein, include the variousisomers such as the α and β isomers (e.g., α-sitosterol andβ-sitostanol, which comprise one of the most effective phytosterols andphytostanols, respectively, for lowering serum cholesterol in mammals).

The phytosterols and phytostanols of the present invention also may bein their ester form. Suitable methods for deriving the esters ofphytosterols and phytostanols are well known to those of ordinary skillin the art, and are disclosed in U.S. Pat. Nos. 6,589,588, 6,635,774,6,800,317, and U.S. Patent Publication Number 2003/0045473, thedisclosures of which are incorporated herein by reference in theirentirety. Non-limiting examples of suitable phytosterol and phytostanolesters include sitosterol acetate, sitosterol oleate, stigmasterololeate, and their corresponding phytostanol esters. The phytosterols andphytostanols of the present invention also may include theirderivatives.

Generally, the amount of functional ingredient in the sweetenercomposition or sweetened composition varies widely depending on theparticular sweetener composition or sweetened composition and thedesired functional ingredient. Those of ordinary skill in the art willreadily acertain the appropriate amount of functional ingredient foreach sweetener composition or sweetened composition.

In one embodiment, a method for preparing a sweetener compositioncomprises combining Reb X and at least one sweetener and/or additiveand/or functional ingredient. In another embodiment, a method forpreparing a sweetener composition comprises combining a compositioncomprising Reb X and at least one sweetener and/or additive and/orfunctional ingredient. Reb X can be provided in its pure form as thesole sweetener in the sweetener composition, or it can be provided aspart of a steviol glycoside mixture of Stevia extract. Any of thesweeteners, additives and functional ingredients described herein can beused the in the sweetener compositions of the present invention.

Sweetened Compositions

Reb X or sweetener compositions comprising Reb X can be incorporated inany known edible material (referred to herein as a “sweetenablecomposition”), such as, for example, pharmaceutical compositions, ediblegel mixes and compositions, dental compositions, foodstuffs(confections, condiments, chewing gum, cereal compositions baked goodsdairy products, and tabletop sweetener compositions) beverages andbeverage products.

In one embodiment, sweetened composition comprises a sweetenablecomposition and Reb X. In another embodiment, the sweetened compositioncomprises a sweetener composition comprising Reb X. The sweetenedcompositions can optionally include additives, sweeteners, functionalingredients and combinations thereof.

In one embodiment, a method for preparing a sweetened compositioncomprises combining a sweetenable composition and Reb X. The method canfurther comprise adding and at least one sweetener and/or additiveand/or functional ingredient. In another embodiment, a method forpreparing a sweetened composition comprises combining a sweetenablecomposition and a sweetener composition comprising Reb X. Reb X can beprovided in its pure form as the sole sweetener in the sweetenercomposition, or it can be provided as part of a steviol glycosidemixture of Stevia extract. Any of the sweeteners, additives andfunctional ingredients described herein can be used the in the sweetenedcompositions of the present invention. In a particular embodiment, thesweetenable composition is a beverage.

Pharmaceutical Compositions

In one embodiment, a pharmaceutical composition contains apharmaceutically active substance and Reb X. In another embodiment, apharmaceutical composition contains a pharmaceutically active substanceand a sweetener composition comprising Reb X. The Reb X or Reb Xsweetener composition can be present as an excipient material in thepharmaceutical composition, which can mask a bitter or otherwiseundesirable taste of a pharmaceutically active substance or anotherexcipient material. The pharmaceutical composition may be in the form ofa tablet, a capsule, a liquid, an aerosol, a powder, an effervescenttablet or powder, a syrup, an emulsion, a suspension, a solution, or anyother form for providing the pharmaceutical composition to a patient. Inparticular embodiments, the pharmaceutical composition may be in a formfor oral administration, buccal administration, sublingualadministration, or any other route of administration as known in theart.

As referred to herein, “pharmaceutically active substance” means anydrug, drug formulation, medication, prophylactic agent, therapeuticagent, or other substance having biological activity. As referred toherein, “excipient material” refers to any inactive substance used as avehicle for an active ingredient, such as any material to facilitatehandling, stability, dispersibility, wettability, and/or releasekinetics of a pharmaceutically active substance.

Suitable pharmaceutically active substances include, but are not limitedto, medications for the gastrointestinal tract or digestive system, forthe cardiovascular system, for the central nervous system, for pain orconsciousness, for musculo-skeletal disorders, for the eye, for the ear,nose and oropharynx, for the respiratory system, for endocrine problems,for the reproductive system or urinary system, for contraception, forobstetrics and gynecology, for the skin, for infections andinfestations, for immunology, for allergic disorders, for nutrition, forneoplastic disorders, for diagnostics, for euthanasia, or otherbiological functions or disorders. Examples of suitable pharmaceuticallyactive substances for embodiments of the present invention include, butare not limited to, antacids, reflux suppressants, antiflatulents,antidopaminergics, proton pump inhibitors, cytoprotectants,prostaglandin analogues, laxatives, antispasmodics, antidiarrhoeals,bile acid sequestrants, opioids, beta-receptor blockers, calcium channelblockers, diuretics, cardiac glycosides, antiarrhythmics, nitrates,antianginals, vasoconstrictors, vasodilators, peripheral activators, ACEinhibitors, angiotensin receptor blockers, alpha blockers,anticoagulants, heparin, antiplatelet drugs, fibrinolytics,anti-hemophilic factors, haemostatic drugs, hypolipidaemic agents,statins, hynoptics, anaesthetics, antipsychotics, antidepressants,anti-emetics, anticonvulsants, antiepileptics, anxiolytics,barbiturates, movement disorder drugs, stimulants, benzodiazepines,cyclopyrrolones, dopamine antagonists, antihistamines, cholinergics,anticholinergics, emetics, cannabinoids, analgesics, muscle relaxants,antibiotics, aminoglycosides, anti-virals, anti-fungals,anti-inflammatories, anti-gluacoma drugs, sympathomimetics, steroids,ceruminolytics, bronchodilators, NSAIDS, antitussive, mucolytics,decongestants, corticosteroids, androgens, antiandrogens, gonadotropins,growth hormones, insulin, antidiabetics, thyroid hormones, calcitonin,diphosponates, vasopressin analogues, alkalizing agents, quinolones,anticholinesterase, sildenafil, oral contraceptives, Hormone ReplacementTherapies, bone regulators, follicle stimulating hormones, luteinizingshormones, gamolenic acid, progestogen, dopamine agonist, oestrogen,prostaglandin, gonadorelin, clomiphene, tamoxifen, diethylstilbestrol,antileprotics, antituberculous drugs, antimalarials, anthelmintics,antiprotozoal, antiserums, vaccines, interferons, tonics, vitamins,cytotoxic drugs, sex hormones, aromatase inhibitors, somatostatininhibitors, or similar type substances, or combinations thereof. Suchcomponents generally are recognized as safe (GRAS) and/or are U.S. Foodand Drug Administration (FDA)-approved.

The pharmaceutically active substance is present in the pharmaceuticalcomposition in widely ranging amounts depending on the particularpharmaceutically active agent being used and its intended applications.An effective dose of any of the herein described pharmaceutically activesubstances can be readily determined by the use of conventionaltechniques and by observing results obtained under analogouscircumstances. In determining the effective dose, a number of factorsare considered including, but not limited to: the species of thepatient; its size, age, and general health; the specific diseaseinvolved; the degree of involvement or the severity of the disease; theresponse of the individual patient; the particular pharmaceuticallyactive agent administered; the mode of administration; thebioavailability characteristic of the preparation administered; the doseregimen selected; and the use of concomitant medication. Thepharmaceutically active substance is included in the pharmaceuticallyacceptable carrier, diluent, or excipient in an amount sufficient todeliver to a patient a therapeutic amount of the pharmaceutically activesubstance in vivo in the absence of serious toxic effects when used ingenerally acceptable amounts. Thus, suitable amounts can be readilydiscerned by those skilled in the art.

According to particular embodiments of the present invention, theconcentration of pharmaceutically active substance in the pharmaceuticalcomposition will depend on absorption, inactivation, and excretion ratesof the drug as well as other factors known to those of skill in the art.It is to be noted that dosage values will also vary with the severity ofthe condition to be alleviated. It is to be further understood that forany particular subject, specific dosage regimes should be adjusted overtime according to the individual need and the professional judgment ofthe person administering or supervising the administration of thepharmaceutical compositions, and that the dosage ranges set forth hereinare exemplary only and are not intended to limit the scope or practiceof the claimed composition. The pharmaceutically active substance may beadministered at once, or may be divided into a number of smaller dosesto be administered at varying intervals of time.

The pharmaceutical composition also may comprise other pharmaceuticallyacceptable excipient materials in addition to Reb X or a sweetenercomposition comprising Reb X. Examples of suitable excipient materialsfor embodiments of this invention include, but are not limited to,antiadherents, binders (e.g., microcrystalline cellulose, gumtragacanth, or gelatin), coatings, disintegrants, fillers, diluents,softeners, emulsifiers, flavoring agents, coloring agents, adjuvants,lubricants, functional agents (e.g., nutrients), viscosity modifiers,bulking agents, glidiants (e.g., colloidal silicon dioxide) surfaceactive agents, osmotic agents, diluents, or any other non-activeingredient, or combinations thereof. For example, the pharmaceuticalcompositions of the present invention may include excipient materialsselected from the group consisting of calcium carbonate, coloringagents, whiteners, preservatives, and flavors, triacetin, magnesiumstearate, sterotes, natural or artificial flavors, essential oils, plantextracts, fruit essences, gelatins, or combinations thereof.

The excipient material of the pharmaceutical composition may optionallyinclude other artificial or natural sweeteners, bulk sweeteners, orcombinations thereof. Bulk sweeteners include both caloric andnon-caloric compounds. In a particular embodiment, the additivefunctions as the bulk sweetener. Non-limiting examples of bulksweeteners include sucrose, dextrose, maltose, dextrin, dried invertsugar, fructose, high fructose corn syrup, levulose, galactose, cornsyrup solids, tagatose, polyols (e.g., sorbitol, mannitol, xylitol,lactitol, erythritol, and maltitol), hydrogenated starch hydrolysates,isomalt, trehalose, and mixtures thereof. In particular embodiments, thebulk sweetener is present in the pharmaceutical composition in widelyranging amounts depending on the degree of sweetness desired. Suitableamounts of both sweeteners would be readily discernable to those skilledin the art.

Edible Gel Mixes and Edible Gel Compositions

In one embodiment, an edible gel or edible gel mix comprises Reb X. Inanother embodiment, an edible gel or edible gel mix comprises asweetener composition comprising Reb X. The edible gel or edible gelmixes can optionally include additives, functional ingredients orcombinations thereof

Edible gels are gels that can be eaten. A gel is a colloidal system inwhich a network of particles spans the volume of a liquid medium.Although gels mainly are composed of liquids, and thus exhibit densitiessimilar to liquids, gels have the structural coherence of solids due tothe network of particles that spans the liquid medium. For this reason,gels generally appear to be solid, jelly-like materials. Gels can beused in a number of applications. For example, gels can be used infoods, paints, and adhesives.

Non-limiting examples of edible gel compositions for use in particularembodiments include gel desserts, puddings, jellies, pastes, trifles,aspics, marshmallows, gummy candies, or the like. Edible gel mixesgenerally are powdered or granular solids to which a fluid may be addedto form an edible gel composition. Non-limiting examples of fluids foruse in particular embodiments include water, dairy fluids, dairyanalogue fluids, juices, alcohol, alcoholic beverages, and combinationsthereof. Non-limiting examples of dairy fluids which may be used inparticular embodiments include milk, cultured milk, cream, fluid whey,and mixtures thereof. Non-limiting examples of dairy analogue fluidswhich may be used in particular embodiments include, for example, soymilk and non-dairy coffee whitener. Because edible gel products found inthe marketplace typically are sweetened with sucrose, it is desirable tosweeten edible gels with an alternative sweetener in order provide alow-calorie or non-calorie alternative.

As used herein, the term “gelling ingredient” denotes any material thatcan form a colloidal system within a liquid medium. Non-limitingexamples of gelling ingredients for use in particular embodimentsinclude gelatin, alginate, carageenan, gum, pectin, konjac, agar, foodacid, rennet, starch, starch derivatives, and combinations thereof. Itis well known to those having ordinary skill in the art that the amountof gelling ingredient used in an edible gel mix or an edible gelcomposition varies considerably depending on a number of factors, suchas the particular gelling ingredient used, the particular fluid baseused, and the desired properties of the gel.

It is well known to those having ordinary skill in the art that theedible gel mixes and edible gels may be prepared using other ingredientsin addition to Reb X, or the sweetener composition comprising Reb X, andthe gelling agent. Non-limiting examples of other ingredients for use inparticular embodiments include a food acid, a salt of a food acid, abuffering system, a bulking agent, a sequestrant, a cross-linking agent,one or more flavors, one or more colors, and combinations thereof.Non-limiting examples of food acids for use in particular embodimentsinclude citric acid, adipic acid, fumaric acid, lactic acid, malic acid,and combinations thereof. Non-limiting examples of salts of food acidsfor use in particular embodiments include sodium salts of food acids,potassium salts of food acids, and combinations thereof. Non-limitingexamples of bulking agents for use in particular embodiments includeraftilose, isomalt, sorbitol, polydextrose, maltodextrin, andcombinations thereof. Non-limiting examples of sequestrants for use inparticular embodiments include calcium disodium ethylene tetra-acetate,glucono delta-lactone, sodium gluconate, potassium gluconate,ethylenediaminetetraacetic acid (EDTA), and combinations thereof.Non-limiting examples of cross-linking agents for use in particularembodiments include calcium ions, magnesium ions, sodium ions, andcombinations thereof.

Dental Compositions

In one embodiment, a dental composition comprises Reb X. In anotherembodiment, a dental composition comprises a sweetener compositioncomprising Reb X. Dental compositions generally comprise an activedental substance and a base material. Reb X, or a sweetener compositioncomprising Reb X, can be used as the base material to sweeten the dentalcomposition. The dental composition may be in the form of any oralcomposition used in the oral cavity such as mouth freshening agents,gargling agents, mouth rinsing agents, toothpaste, tooth polish,dentifrices, mouth sprays, teeth-whitening agent, dental floss, and thelike, for example.

As referred to herein, “active dental substance” means any compositionwhich can be used to improve the aesthetic appearance and/or health ofteeth or gums or prevent dental caries. As referred to herein, “basematerial” refers to any inactive substance used as a vehicle for anactive dental substance, such as any material to facilitate handling,stability, dispersibility, wettability, foaming, and/or release kineticsof an active dental substance.

Suitable active dental substances for embodiments of this inventioninclude, but are not limited to, substances which remove dental plaque,remove food from teeth, aid in the elimination and/or masking ofhalitosis, prevent tooth decay, and prevent gum disease (i.e., Gingiva).Examples of suitable active dental substances for embodiments of thepresent invention include, but are not limited to, anticaries drugs,fluoride, sodium fluoride, sodium monofluorophosphate, stannos fluoride,hydrogen peroxide, carbamide peroxide (i.e., urea peroxide),antibacterial agents, plaque removing agents, stain removers,anticalculus agents, abrasives, baking soda, percarbonates, perboratesof alkali and alkaline earth metals, or similar type substances, orcombinations thereof. Such components generally are recognized as safe(GRAS) and/or are U.S. Food and Drug Administration (FDA)-approved.

According to particular embodiments of the invention, the active dentalsubstance is present in the dental composition in an amount ranging fromabout 50 ppm to about 3000 ppm of the dental composition. Generally, theactive dental substance is present in the dental composition in anamount effective to at least improve the aesthetic appearance and/orhealth of teeth or gums marginally or prevent dental caries. Forexample, a dental composition comprising a toothpaste may include anactive dental substance comprising fluoride in an amount of about 850 to1,150 ppm.

The dental composition also may comprise other base materials inaddition to the Reb X or sweetener composition comprising Reb X.Examples of suitable base materials for embodiments of this inventioninclude, but are not limited to, water, sodium lauryl sulfate or othersulfates, humectants, enzymes, vitamins, herbs, calcium, flavorings(e.g., mint, bubblegum, cinnamon, lemon, or orange), surface-activeagents, binders, preservatives, gelling agents, pH modifiers, peroxideactivators, stabilizers, coloring agents, or similar type materials, andcombinations thereof.

The base material of the dental composition may optionally include otherartificial or natural sweeteners, bulk sweeteners, or combinationsthereof. Bulk sweeteners include both caloric and non-caloric compounds.Non-limiting examples of bulk sweeteners include sucrose, dextrose,maltose, dextrin, dried invert sugar, fructose, high fructose cornsyrup, levulose, galactose, corn syrup solids, tagatose, polyols (e.g.,sorbitol, mannitol, xylitol, lactitol, erythritol, and maltitol),hydrogenated starch hydrolysates, isomalt, trehalose, and mixturesthereof. Generally, the amount of bulk sweetener present in the dentalcomposition ranges widely depending on the particular embodiment of thedental composition and the desired degree of sweetness. Those ofordinary skill in the art will readily ascertain the appropriate amountof bulk sweetener. In particular embodiments, the bulk sweetener ispresent in the dental composition in an amount in the range of about 0.1to about 5 weight percent of the dental composition.

According to particular embodiments of the invention, the base materialis present in the dental composition in an amount ranging from about 20to about 99 percent by weight of the dental composition. Generally, thebase material is present in an amount effective to provide a vehicle foran active dental substance.

In a particular embodiment, a dental composition comprises Reb X and anactive dental substance. In another particular embodiment, a dentalcomposition comprises a sweetener composition comprising Reb X and anactive dental substance. Generally, the amount of the sweetener varieswidely depending on the nature of the particular dental composition andthe desired degree of sweetness. Those skilled in the art will be ableto discern a suitable amount of sweetener for such dental composition.In a particular embodiment, Reb X is present in the dental compositionin an amount in the range of about 1 to about 5,000 ppm of the dentalcomposition and the at least one additive is present in the dentalcomposition in an amount in the range of about 0.1 to about 100,000 ppmof the dental composition.

Foodstuffs include, but are not limited to, confections, condiments,chewing gum, cereal, baked goods, and dairy products.

Confections

In one embodiment, a confection comprises Reb X. In another embodiment,a confection comprises a sweetener composition comprising Reb X.

As referred to herein, “confection” can mean a sweet, a lollie, aconfectionery, or similar term. The confection generally contains a basecomposition component and a sweetener component. Reb X or a sweetenercomposition comprising Reb X can serve as the sweetener component. Theconfection may be in the form of any food that is typically perceived tobe rich in sugar or is typically sweet. According to particularembodiments of the present invention, the confections may be bakeryproducts such as pastries; desserts such as yogurt, jellies, drinkablejellies, puddings, Bavarian cream, blancmange, cakes, brownies, mousseand the like, sweetened food products eaten at tea time or followingmeals; frozen foods; cold confections, e. g. types of ice cream such asice cream, ice milk, lacto-ice and the like (food products in whichsweeteners and various other types of raw materials are added to milkproducts, and the resulting mixture is agitated and frozen), and iceconfections such as sherbets, dessert ices and the like (food productsin which various other types of raw materials are added to a sugaryliquid, and the resulting mixture is agitated and frozen); generalconfections, e. g., baked confections or steamed confections such ascrackers, biscuits, buns with bean-jam filling, halvah, alfajor, and thelike; rice cakes and snacks; table top products; general sugarconfections such as chewing gum (e.g. including compositions whichcomprise a substantially water-insoluble, chewable gum base, such aschicle or substitutes thereof, including jetulong, guttakay rubber orcertain comestible natural synthetic resins or waxes), hard candy, softcandy, mints, nougat candy, jelly beans, fudge, toffee, taffy, Swissmilk tablet, licorice candy, chocolates, gelatin candies, marshmallow,marzipan, divinity, cotton candy, and the like; sauces including fruitflavored sauces, chocolate sauces and the like; edible gels; crèmesincluding butter crèmes, flour pastes, whipped cream and the like; jamsincluding strawberry jam, marmalade and the like; and breads includingsweet breads and the like or other starch products, and combinationsthereof.

As referred to herein, “base composition” means any composition whichcan be a food item and provides a matrix for carrying the sweetenercomponent.

Suitable base compositions for embodiments of this invention may includeflour, yeast, water, salt, butter, eggs, milk, milk powder, liquor,gelatin, nuts, chocolate, citric acid, tartaric acid, fumaric acid,natural flavors, artificial flavors, colorings, polyols, sorbitol,isomalt, maltitol, lactitol, malic acid, magnesium stearate, lecithin,hydrogenated glucose syrup, glycerine, natural or synthetic gum, starch,and the like, and combinations thereof. Such components generally arerecognized as safe (GRAS) and/or are U.S. Food and Drug Administration(FDA)-approved. According to particular embodiments of the invention,the base composition is present in the confection in an amount rangingfrom about 0.1 to about 99 weight percent of the confection. Generally,the base composition is present in the confection in an amount, incombination with Reb X or a sweetener composition comprising Reb X toprovide a food product.

The base composition of the confection may optionally include otherartificial or natural sweeteners, bulk sweeteners, or combinationsthereof. Bulk sweeteners include both caloric and non-caloric compounds.Non-limiting examples of bulk sweeteners include sucrose, dextrose,maltose, dextrin, dried invert sugar, fructose, high fructose cornsyrup, levulose, galactose, corn syrup solids, tagatose, polyols (e.g.,sorbitol, mannitol, xylitol, lactitol, erythritol, and maltitol),hydrogenated starch hydrolysates, isomalt, trehalose, and mixturesthereof. Generally, the amount of bulk sweetener present in theconfection ranges widely depending on the particular embodiment of theconfection and the desired degree of sweetness. Those of ordinary skillin the art will readily ascertain the appropriate amount of bulksweetener.

In a particular embodiment, a confection comprises Reb X, or a sweetenercomposition comprising Reb X, and a base composition. Generally, theamount of Reb X in the confection ranges widely depending on theparticular embodiment of the confection and the desired degree ofsweetness. Those of ordinary skill in the art will readily ascertain theappropriate amount of sweetener. In a particular embodiment, Reb X ispresent in the confection in an amount in the range of about 30 ppm toabout 6000 ppm of the confection. In another embodiment, Reb X ispresent in the confection in an amount in the range of about 1 ppm toabout 10,000 ppm of the confection. In embodiments where the confectioncomprises hard candy, Reb X is present in an amount in the range ofabout 150 ppm to about 2250 ppm of the hard candy.

Condiment Compositions

In one embodiment, a condiment comprises Reb X. In another embodiment acondiment comprises a sweetener composition comprising Reb X.Condiments, as used herein, are compositions used to enhance or improvethe flavor of a food or beverage. Non-limiting examples of condimentsinclude ketchup (catsup); mustard; barbecue sauce; butter; chili sauce;chutney; cocktail sauce; curry; dips; fish sauce; horseradish; hotsauce; jellies, jams, marmalades, or preserves; mayonnaise; peanutbutter; relish; remoulade; salad dressings (e.g., oil and vinegar,Caesar, French, ranch, bleu cheese, Russian, Thousand Island, Italian,and balsamic vinaigrette), salsa; sauerkraut; soy sauce; steak sauce;syrups; tartar sauce; and Worcestershire sauce.

Condiment bases generally comprise a mixture of different ingredients,non-limiting examples of which include vehicles (e.g., water andvinegar); spices or seasonings (e.g., salt, pepper, garlic, mustardseed, onion, paprika, turmeric, and combinations thereof); fruits,vegetables, or their products (e.g., tomatoes or tomato-based products(paste, puree), fruit juices, fruit juice peels, and combinationsthereof); oils or oil emulsions, particularly vegetable oils; thickeners(e.g., xanthan gum, food starch, other hydrocolloids, and combinationsthereof); and emulsifying agents (e.g., egg yolk solids, protein, gumarabic, carob bean gum, guar gum, gum karaya, gum tragacanth,carageenan, pectin, propylene glycol esters of alginic acid, sodiumcarboxymethyl-cellulose, polysorbates, and combinations thereof).Recipes for condiment bases and methods of making condiment bases arewell known to those of ordinary skill in the art.

Generally, condiments also comprise caloric sweeteners, such as sucrose,high fructose corn syrup, molasses, honey, or brown sugar. In exemplaryembodiments of the condiments provided herein, Reb X or sweetenercompositions comprising Reb X is used instead of traditional caloricsweeteners. Accordingly, a condiment composition desirably comprises RebX or a sweetener composition comprising Reb X and a condiment base.

The condiment composition optionally may include other natural and/orsynthetic high-potency sweeteners, bulk sweeteners, pH modifying agents(e.g., lactic acid, citric acid, phosphoric acid, hydrochloric acid,acetic acid, and combinations thereof), fillers, functional agents(e.g., pharmaceutical agents, nutrients, or components of a food orplant), flavorings, colorings, or combinations thereof.

Chewing Gum Compositions

In one embodiment, a chewing gum composition comprises Reb X. In anotherembodiment, a chewing gum composition comprises a sweetener compositioncomprising Reb X. Chewing gum compositions generally comprise awater-soluble portion and a water-insoluble chewable gum base portion.The water soluble portion, which typically includes the sweetener orsweetener composition, dissipates with a portion of the flavoring agentover a period of time during chewing while the insoluble gum baseportion is retained in the mouth. The insoluble gum base generallydetermines whether a gum is considered chewing gum, bubble gum, or afunctional gum.

The insoluble gum base, which is generally present in the chewing gumcomposition in an amount in the range of about 15 to about 35 weightpercent of the chewing gum composition, generally comprises combinationsof elastomers, softeners (plasticizers), emulsifiers, resins, andfillers. Such components generally are considered food grade, recognizedas safe (GRA), and/or are U.S. Food and Drug Administration(FDA)-approved.

Elastomers, the primary component of the gum base, provide the rubbery,cohesive nature to gums and can include one or more natural rubbers(e.g., smoked latex, liquid latex, or guayule); natural gums (e.g.,jelutong, perillo, sorva, massaranduba balata, massaranduba chocolate,nispero, rosindinha, chicle, and gutta hang kang); or syntheticelastomers (e.g., butadiene-styrene copolymers, isobutylene-isoprenecopolymers, polybutadiene, polyisobutylene, and vinyl polymericelastomers). In a particular embodiment, the elastomer is present in thegum base in an amount in the range of about 3 to about 50 weight percentof the gum base.

Resins are used to vary the firmness of the gum base and aid insoftening the elastomer component of the gum base. Non-limiting examplesof suitable resins include a rosin ester, a terpene resin (e.g., aterpene resin from α-pinene, β-pinene and/or d-limonene), polyvinylacetate, polyvinyl alcohol, ethylene vinyl acetate, and vinylacetate-vinyl laurate copolymers. Non-limiting examples of rosin estersinclude a glycerol ester of a partially hydrogenated rosin, a glycerolester of a polymerized rosin, a glycerol ester of a partially dimerizedrosin, a glycerol ester of rosin, a pentaerythritol ester of a partiallyhydrogenated rosin, a methyl ester of rosin, or a methyl ester of apartially hydrogenated rosin. In a particular embodiment, the resin ispresent in the gum base in an amount in the range of about 5 to about 75weight percent of the gum base.

Softeners, which also are known as plasticizers, are used to modify theease of chewing and/or mouthfeel of the chewing gum composition.Generally, softeners comprise oils, fats, waxes, and emulsifiers,Non-limiting examples of oils and fats include tallow, hydrogenatedtallow, large, hydrogenated or partially hydrogenated vegetable oils(e.g., soybean, canola, cottonseed, sunflower, palm, coconut, corn,safflower, or palm kernel oils), cocoa butter, glycerol monostearate,glycerol triacetate, glycerol abietate, leithin, monoglycerides,diglycerides, triglycerides acetylated monoglycerides, and free fattyacids. Non-limiting examples of waxes includepolypropylene/polyethylene/Fisher-Tropsch waxes, paraffin, andmicrocrystalline and natural waxes (e.g., candelilla, beeswas andcarnauba). Microcrystalline waxes, especially those with a high degreeof crystallinity and a high melting point, also may be considered asbodying agents or textural modifiers. In a particular embodiment, thesofteners are present in the gum base in an amount in the range of about0.5 to about 25 weight percent of the gum base.

Emulsifiers are used to form a uniform dispersion of the insoluble andsoluble phases of the chewing gum composition and also have plasticizingproperties. Suitable emulsifiers include glycerol monostearate (GMS),lecithin (Phosphatidyl choline), polyglycerol polyricinoleic acid(PPGR), mono and diglycerides of fatty acids, glycerol distearate,tracetin, acetylated monoglyceride, glycerol triactetate, and magnesiumstearate. In a particular embodiment, the emulsifiers are present in thegum base in an amount in the range of about 2 to about 30 weight percentof the gum base.

The chewing gum composition also may comprise adjuvants or fillers ineither the gum base and/or the soluble portion of the chewing gumcomposition. Suitable adjuvants and fillers include lecithin, inulin,polydextrin, calcium carbonate, magnesium carbonate, magnesium silicate,ground limestome, aluminum hydroxide, aluminum silicate, talc, clay,alumina, titanium dioxide, and calcium phosphate. In particularembodiments, lecithin can be used as an inert filler to decrease thestickiness of the chewing gum composition. In other particularembodiments, lactic acid copolymers, proteins (e.g., gluten and/or zein)and/or guar can be used to create a gum that is more readilybiodegradable. The adjuvants or fillers are generally present in the gumbase in an amount up to about 20 weight percent of the gum base. Otheroptional ingredients include coloring agents, whiteners, preservatives,and flavors.

In particular embodiments of the chewing gum composition, the gum basecomprises about 5 to about 95 weight percent of the chewing gumcomposition, more desirably about 15 to about 50 weight percent of thechewing gum composition, and even more desirably from about 20 to about30 weight percent of the chewing gum composition.

The soluble portion of the chewing gum composition may optionallyinclude other artificial or natural sweeteners, bulk sweeteners,softeners, emulsifiers, flavoring agents, coloring agents, adjuvants,fillers, functional agents (e.g., pharmaceutical agents or nutrients),or combinations thereof. Suitable examples of softeners and emulsifiersare described above.

Bulk sweeteners include both caloric and non-caloric compounds.Non-limiting examples of bulk sweeteners include sucrose, dextrose,maltose, dextrin, dried invert sugar, fructose, high fructose cornsyrup, levulose, galactose, corn syrup solids, tagatose, polyols (e.g.,sorbitol, mannitol, xylitol, lactitol, erythritol, and maltitol),hydrogenated starch hydrolysates, isomalt, trehalose, and mixturesthereof. In particular embodiments, the bulk sweetener is present in thechewing gum composition in an amount in the range of about 1 to about 75weight percent of the chewing gum composition.

Flavoring agents may be used in either the insoluble gum base or solubleportion of the chewing gum composition. Such flavoring agents may benatural or artificial flavors. In a particular embodiment, the flavoringagent comprises an essential oil, such as an oil derived from a plant ora fruit, peppermint oil, spearmint oil, other mint oils, clove oil,cinnamon oil, oil of wintergreen, bay, thyme, cedar leaf, nutmeg,allspice, sage, mace, and almonds. In another particular embodiment, theflavoring agent comprises a plant extract or a fruit essence such asapple, banana, watermelon, pear, peach, grape, strawberry, raspberry,cherry, plum, pineapple, apricot, and mixtures thereof. In still anotherparticular embodiment, the flavoring agent comprises a citrus flavor,such as an extract, essence, or oil of lemon, lime, orange, tangerine,grapefruit, citron, or kumquat.

In a particular embodiment, a chewing gum composition comprises or asweetener composition comprising Reb X and a gum base. In a particularembodiment, Reb X is present in the chewing gum composition in an amountin the range of about 1 ppm to about 10,000 ppm of the chewing gumcomposition.

Cereal Compositions

In one embodiment, a cereal composition comprises Reb X. In anotherembodiment, a cereal composition comprises a sweetener compositioncomprising Reb X. Cereal compositions typically are eaten either asstaple foods or as snacks. Non-limiting examples of cereal compositionsfor use in particular embodiments include ready-to-eat cereals as wellas hot cereals. Ready-to-eat cereals are cereals which may be eatenwithout further processing (i.e. cooking) by the consumer. Examples ofready-to-eat cereals include breakfast cereals and snack bars. Breakfastcereals typically are processed to produce a shredded, flaky, puffy, orextruded form. Breakfast cereals generally are eaten cold and are oftenmixed with milk and/or fruit. Snack bars include, for example, energybars, rice cakes, granola bars, and nutritional bars. Hot cerealsgenerally are cooked, usually in either milk or water, before beingeaten. Non-limiting examples of hot cereals include grits, porridge,polenta, rice, and rolled oats.

Cereal compositions generally comprise at least one cereal ingredient.As used herein, the term “cereal ingredient” denotes materials such aswhole or part grains, whole or part seeds, and whole or part grass.Non-limiting examples of cereal ingredients for use in particularembodiments include maize, wheat, rice, barley, bran, bran endosperm,bulgur, soghums, millets, oats, rye, triticale, buchwheat, fonio,quinoa, bean, soybean, amaranth, teff, spelt, and kaniwa.

In a particular embodiment, the cereal composition comprises Reb X or asweetener composition comprising Reb X and at least one cerealingredient. Reb X or the sweetener composition comprising Reb X may beadded to the cereal composition in a variety of ways, such as, forexample, as a coating, as a frosting, as a glaze, or as a matrix blend(i.e. added as an ingredient to the cereal formulation prior to thepreparation of the final cereal product).

Accordingly, in a particular embodiment, Reb X or a sweetenercomposition comprising Reb X is added to the cereal composition as amatrix blend. In one embodiment, Reb X or a sweetener compositioncomprising Reb X is blended with a hot cereal prior to cooking toprovide a sweetened hot cereal product. In another embodiment, Reb X ora sweetener comprising Reb X is blended with the cereal matrix beforethe cereal is extruded.

In another particular embodiment, Reb X or a sweetener compositioncomprising Reb X is added to the cereal composition as a coating, suchas, for example, by combining Reb X or a sweetener comprising Reb X witha food grade oil and applying the mixture onto the cereal. In adifferent embodiment, Reb X or a sweetener composition comprising Reb Xand the food grade oil may be applied to the cereal separately, byapplying either the oil or the sweetener first. Non-limiting examples offood grade oils for use in particular embodiments include vegetable oilssuch as corn oil, soybean oil, cottonseed oil, peanut oil, coconut oil,canola oil, olive oil, sesame seed oil, palm oil, palm kernel oil, andmixtures thereof. In yet another embodiment, food grade fats may be usedin place of the oils, provided that the fat is melted prior to applyingthe fat onto the cereal.

In another embodiment, the Reb X or a sweetener composition comprisingReb X is added to the cereal composition as a glaze. Non-limitingexamples of glazing agents for use in particular embodiments includecorn syrup, honey syrups and honey syrup solids, maple syrups and maplesyrup solids, sucrose, isomalt, polydextrose, polyols, hydrogenatedstarch hydrosylate, aqueous solutions thereof, and mixtures thereof. Inanother such embodiment, Reb X or a sweetener composition comprising RebX is added as a glaze by combining with a glazing agent and a food gradeoil or fat and applying the mixture to the cereal. In yet anotherembodiment, a gum system, such as, for example, gum acacia,carboxymethyl cellulose, or algin, may be added to the glaze to providestructural support. In addition, the glaze also may include a coloringagent, and also may include a flavor.

In another embodiment, Reb X or a sweetener composition comprising Reb Xis added to the cereal composition as a frosting. In one suchembodiment, Reb X or a sweetener composition comprising Reb X iscombined with water and a frosting agent and then applied to the cereal.Non-limiting examples of frosting agents for use in particularembodiments include maltodextrin, sucrose, starch, polyols, and mixturesthereof. The frosting also may include a food grade oil, a food gradefat, a coloring agent, and/or a flavor.

Generally, the amount of Reb X in a cereal composition varies widelydepending on the particular type of cereal composition and its desiredsweetness. Those of ordinary skill in the art can readily discern theappropriate amount of sweetener to put in the cereal composition. In aparticular embodiment, Reb X is present in the cereal composition in anamount in the range of about 0.02 to about 1.5 weight percent of thecereal composition and the at least one additive is present in thecereal composition in an amount in the range of about 1 to about 5weight percent of the cereal composition.

Baked Goods

In one embodiment, a baked good comprises Reb X. In another embodiment,a baked good comprises a sweetener composition comprising Reb X. Bakedgoods, as used herein, include ready to eat and all ready to bakeproducts, flours, and mixes requiring preparation before serving.Non-limiting examples of baked goods include cakes, crackers, cookies,brownies, muffins, rolls, bagels, donuts, strudels, pastries,croissants, biscuits, bread, bread products, and buns.

Preferred baked goods in accordance with embodiments of this inventioncan be classified into three groups: bread-type doughs (e.g., whitebreads, variety breads, soft buns, hard rolls, bagels, pizza dough, andflour tortillas), sweet doughs (e.g., danishes, croissants, crackers,puff pastry, pie crust, biscuits, and cookies), and batters (e.g., cakessuch as sponge, pound, devil's food, cheesecake, and layer cake, donutsor other yeast raised cakes, brownies, and muffins). Doughs generallyare characterized as being flour-based, whereas batters are morewater-based.

Baked goods in accordance with particular embodiments of this inventiongenerally comprise a combination of sweetener, water, and fat. Bakedgoods made in accordance with many embodiments of this invention alsocontain flour in order to make a dough or a batter. The term “dough” asused herein is a mixture of flour and other ingredients stiff enough toknead or roll. The term “batter” as used herein consists of flour,liquids such as milk or water, and other ingredients, and is thin enoughto pour or drop from a spoon. Desirably, in accordance with particularembodiments of the invention, the flour is present in the baked goods inan amount in the range of about 15 to about 60% on a dry weight basis,more desirably from about 23 to about 48% on a dry weight basis.

The type of flour may be selected based on the desired product.Generally, the flour comprises an edible non-toxic flour that isconventionally utilized in baked goods. According to particularembodiments, the flour may be a bleached bake flour, general purposeflour, or unbleached flour. In other particular embodiments, flours alsomay be used that have been treated in other manners. For example, inparticular embodiments flour may be enriched with additional vitamins,minerals, or proteins. Non-limiting examples of flours suitable for usein particular embodiments of the invention include wheat, corn meal,whole grain, fractions of whole grains (wheat, bran, and oatmeal), andcombinations thereof. Starches or farinaceous material also may be usedas the flour in particular embodiments. Common food starches generallyare derived from potato, corn, wheat, barley, oat, tapioca, arrow root,and sago. Modified starches and pregelatinized starches also may be usedin particular embodiments of the invention.

The type of fat or oil used in particular embodiments of the inventionmay comprise any edible fat, oil, or combination thereof that issuitable for baking. Non-limiting examples of fats suitable for use inparticular embodiments of the invention include vegetable oils, tallow,lard, marine oils, and combinations thereof. According to particularembodiments, the fats may be fractionated, partially hydrogenated,and/or interesterified. In another particular embodiment, the fatdesirably comprises reduced, low calorie, or non-digestible fats, fatsubstitutes, or synthetic fats. In yet another particular embodiment,shortenings, fats, or mixtures of hard and soft fats also may be used.In particular embodiments, shortenings may be derived principally fromtriglycerides derived from vegetable sources (e.g., cotton seed oil,soybean oil, peanut oil, linseed oil, sesame oil, palm oil, palm kerneloil, rapeseed oil, safflower oil, coconut oil, corn oil, sunflower seedoil, and mixtures thereof). Synthetic or natural triglycerides of fattyacids having chain lengths from 8 to 24 carbon atoms also may be used inparticular embodiments. Desirably, in accordance with particularembodiments of this invention, the fat is present in the baked good inan amount in the range of about 2 to about 35% by weight on a dry basis,more desirably from about 3 to about 29% by weight on a dry basis.

Baked goods in accordance with particular embodiments of this inventionalso comprise water in amounts sufficient to provide the desiredconsistency, enabling proper forming, machining and cutting of the bakedgood prior or subsequent to cooking. The total moisture content of thebaked good includes any water added directly to the baked good as wellas water present in separately added ingredients (e.g., flour, whichgenerally includes about 12 to about 14% by weight moisture). Desirably,in accordance with particular embodiments of this invention, the wateris present in the baked good in an amount up to about 25% by weight ofthe baked good.

Baked goods in accordance with particular embodiments of this inventionalso may comprise a number of additional conventional ingredients suchas leavening agents, flavors, colors, milk, milk by-products, egg, eggby-products, cocoa, vanilla or other flavoring, as well as inclusionssuch as nuts, raisins, cherries, apples, apricots, peaches, otherfruits, citrus peel, preservative, coconuts, flavored chips such achocolate chips, butterscotch chips, and caramel chips, and combinationsthereof. In particular embodiments, the baked goods may also compriseemulsifiers, such as lecithin and monoglycerides.

According to particular embodiments of this invention, leavening agentsmay comprise chemical leavening agents or yeast leavening agents.Non-limiting examples of chemical leavening agents suitable for use inparticular embodiments of this invention include baking soda (e.g.,sodium, potassium, or aluminum bicarbonate), baking acid (e.g., sodiumaluminum phosphate, monocalcium phosphate, or dicalcium phosphate), andcombinations thereof.

In accordance with another particular embodiment of this invention,cocoa may comprise natural or “Dutched” chocolate from which asubstantial portion of the fat or cocoa butter has been expressed orremoved by solvent extraction, pressing, or other means. In a particularembodiment, it may be necessary to reduce the amount of fat in a bakedgood comprising chocolate because of the additional fat present in cocoabutter. In particular embodiments, it may be necessary to add largeramounts of chocolate as compared to cocoa in order to provide anequivalent amount of flavoring and coloring.

Baked goods generally also comprise caloric sweeteners, such as sucrose,high fructose corn syrup, erythritol, molasses, honey, or brown sugar.In exemplary embodiments of the baked goods provided herein, the caloricsweetener is replaced partially or totally with Reb X or a sweetenercomposition comprising Reb X. Accordingly, in one embodiment a bakedgood comprises Reb X or a sweetener composition comprising Reb X incombination with a fat, water, and optionally flour. In a particularembodiment, the baked good optionally may include other natural and/orsynthetic high-potency sweeteners and/or bulk sweeteners.

Dairy Products

In one embodiment, a dairy product comprises Reb X. In anotherembodiment, a dairy product comprises a sweetener composition comprisingReb X. Dairy products and processes for making dairy products suitablefor use in this invention are well known to those of ordinary skill inthe art. Daily products, as used herein, comprise milk or foodstuffsproduced from milk. Non-limiting examples of dairy products suitable foruse in embodiments of this invention include milk, milk cream, sourcream, crème fraiche, buttermilk, cultured buttermilk, milk powder,condensed milk, evaporated milk, butter, cheese, cottage cheese, creamcheese, yogurt, ice cream, frozen custard, frozen yogurt, gelato, vla,piima, filmjölk, kajmak, kephir, viili, kumiss, airag, ice milk, casein,ayran, lassi, khoa, or combinations thereof.

Milk is a fluid secreted by the mammary glands of female mammals for thenourishment of their young. The female ability to produce milk is one ofthe defining characteristics of mammals and provides the primary sourceof nutrition for newborns before they are able to digest more diversefoods. In particular embodiments of this invention, the dairy productsare derived from the raw milk of cows, goats, sheep, horses, donkeys,camels, water buffalo, yaks, reindeer, moose, or humans.

In particular embodiments of this invention, the processing of the dairyproduct from raw milk generally comprises the steps of pasteurizing,creaming, and homogenizing. Although raw milk may be consumed withoutpasteurization, it usually is pasteurized to destroy harmfulmicroorganisms such as bacteria, viruses, protozoa, molds, and yeasts.Pasteurizing generally comprises heating the milk to a high temperaturefor a short period of time to substantially reduce the number ofmicroorganisms, thereby reducing the risk of disease.

Creaming traditionally follows pasteurization step, and involves theseparation of milk into a higher-fat cream layer and a lower-fat milklayer. Milk will separate into milk and cream layers upon standing fortwelve to twenty-four hours. The cream rises to the top of the milklayer and may be skimmed and used as a separate dairy product.Alternatively, centrifuges may be used to separate the cream from themilk. The remaining milk is classified according to the fat content ofthe milk, non-limiting examples of which include whole, 2%, 1%, and skimmilk.

After removing the desired amount of fat from the milk by creaming, milkis often homogenized. Homogenization prevents cream from separating fromthe milk and generally involves pumping the milk at high pressuresthrough narrow tubes in order to break up fat globules in the milk.Pasteurization, creaming, and homogenization of milk are common but arenot required to produce consumable dairy products. Accordingly, suitabledairy products for use in embodiments of this invention may undergo noprocessing steps, a single processing step, or combinations of theprocessing steps described herein. Suitable dairy products for use inembodiments of this invention may also undergo processing steps inaddition to or apart from the processing steps described herein.

Particular embodiments of this invention comprise dairy productsproduced from milk by additional processing steps. As described above,cream may be skimmed from the top of milk or separated from the milkusing machine-centrifuges. In a particular embodiment, the dairy productcomprises sour cream, a dairy product rich in fats that is obtained byfermenting cream using a bacterial culture. The bacteria produce lacticacid during fermentation, which sours and thickens the cream. In anotherparticular embodiment, the dairy product comprises crème fraiche, aheavy cream slightly soured with bacterial culture in a similar mannerto sour cream. Crème fraiche ordinarily is not as thick or as sour assour cream. In yet another particular embodiment, the dairy productcomprises cultured buttermilk. Cultured buttermilk is obtained by addingbacteria to milk. The resulting fermentation, in which the bacterialculture turns lactose into lactic acid, gives cultured buttermilk a sourtaste. Although it is produced in a different manner, culturedbuttermilk generally is similar to traditional buttermilk, which is aby-product of butter manufacture.

According to other particular embodiments of this invention, the dairyproducts comprise milk powder, condensed milk, evaporated milk, orcombinations thereof. Milk powder, condensed milk, and evaporated milkgenerally are produced by removing water from milk. In a particularembodiment, the dairy product comprises a milk powder comprising driedmilk solids with a low moisture content. In another particularembodiment, the dairy product comprises condensed milk. Condensed milkgenerally comprises milk with a reduced water content and addedsweetener, yielding a thick, sweet product with a long shelf-life. Inyet another particular embodiment, the dairy product comprisesevaporated milk. Evaporated milk generally comprises fresh, homogenizedmilk from which about 60% of the water has been removed, that has beenchilled, fortified with additives such as vitamins and stabilizers,packaged, and finally sterilized. According to another particularembodiment of this invention, the dairy product comprises a dry creamerand Reb X or a Reb X sweetener composition.

In another particular embodiment, the dairy product provided hereincomprises butter. Butter generally is made by churning fresh orfermented cream or milk. Butter generally comprises butterfatsurrounding small droplets comprising mostly water and milk proteins.The churning process damages the membranes surrounding the microscopicglobules of butterfat, allowing the milk fats to conjoin and to separatefrom the other parts of the cream. In yet another particular embodiment,the dairy product comprises buttermilk, which is the sour-tasting liquidremaining after producing butter from full-cream milk by the churningprocess.

In still another particular embodiment, the dairy product comprisescheese, a solid foodstuff produced by curdling milk using a combinationof rennet or rennet substitutes and acidification. Rennet, a naturalcomplex of enzymes produced in mammalian stomachs to digest milk, isused in cheese-making to curdle the milk, causing it to separate intosolids known as curds and liquids known as whey. Generally, rennet isobtained from the stomachs of young ruminants, such as calves; however,alternative sources of rennet include some plants, microbial organisms,and genetically modified bacteria, fungus, or yeast. In addition, milkmay be coagulated by adding acid, such as citric acid. Generally, acombination of rennet and/or acidification is used to curdle the milk.After separating the milk into curds and whey, some cheeses are made bysimply draining, salting, and packaging the curds. For most cheeses,however, more processing is needed. Many different methods may be usedto produce the hundreds of available varieties of cheese. Processingmethods include heating the cheese, cutting it into small cubes todrain, salting, stretching, cheddaring, washing, molding, aging, andripening. Some cheeses, such as the blue cheeses, have additionalbacteria or molds introduced to them before or during aging, impartingflavor and aroma to the finished product. Cottage cheese is a cheesecurd product with a mild flavor that is drained but not pressed so thatsome whey remains. The curd is usually washed to remove acidity. Creamcheese is a soft, mild-tasting, white cheese with a high fat contentthat is produced by adding cream to milk and then curdling to form arich curd. Alternatively, cream cheese can be made from skim milk withcream added to the curd. It should be understood that cheese, as usedherein, comprises all solid foodstuff produced by the curdling milk.

In another particular embodiment of this invention, the dairy productcomprises yogurt. Yogurt generally is produced by the bacterialfermentation of milk. The fermentation of lactose produces lactic acid,which acts on proteins in milk to give the yogurt a gel-like texture andtartness. In particularly desirable embodiments, the yogurt may besweetened with a sweetener and/or flavored. Non-limiting examples offlavorings include, but are not limited to, fruits (e.g., peach,strawberry, banana), vanilla, and chocolate. Yogurt, as used herein,also includes yogurt varieties with different consistencies andviscosities, such as dahi, dadih or dadiah, labneh or labaneh,bulgarian, kefir, and matsoni. In another particular embodiment, thedairy product comprises a yogurt-based beverage, also known as drinkableyogurt or a yogurt smoothie. In particularly desirable embodiments, theyogurt-based beverage may comprise sweeteners, flavorings, otheringredients, or combinations thereof.

Other dairy products beyond those described herein may be used inparticular embodiments of this invention. Such dairy products are wellknown to those of ordinary skill in the art, non-limiting examples ofwhich include milk, milk and juice, coffee, tea, vla, piima, filmjolk,kajmak, kephir, viili, kumiss, airag, ice milk, casein, ayran, lassi,and khoa.

According to particular embodiments of this invention, the dairycompositions also may comprise other additives. Non-limiting examples ofsuitable additives include sweeteners and flavorants such as chocolate,strawberry, and banana. Particular embodiments of the dairy compositionsprovided herein also may comprise additional nutritional supplementssuch as vitamins (e.g., vitamin D) and minerals (e.g., calcium) toimprove the nutritional composition of the milk.

In a particularly desirable embodiment, the dairy composition comprisesReb X or a sweetener composition comprising Reb X in combination with adairy product. In a particular embodiment, Reb X is present in the dairycomposition in an amount in the range of about 200 to about 20,000weight percent of the dairy composition.

Reb X or sweetener compositions comprising Reb X are also suitable foruse in processed agricultural products, livestock products or seafood;processed meat products such as sausage and the like; retort foodproducts, pickles, preserves boiled in soy sauce, delicacies, sidedishes; soups; snacks such as potato chips, cookies, or the like; asshredded filler, leaf, stem, stalk, homogenized leaf cured and animalfeed.

Tabletop Sweetener Compositions

Tabletop sweetener compositions containing Reb X are also contemplatedherein. The tabletop composition can further include at least onebulking agent, additive, anti-caking agent, functional ingredient orcombination thereof.

Suitable “bulking agents” include, but are not limited to, maltodextrin(10 DE, 18 DE, or 5 DE), corn syrup solids (20 or 36 DE), sucrose,fructose, glucose, invert sugar, sorbitol, xylose, ribulose, mannose,xylitol, mannitol, galactitol, erythritol, maltitol, lactitol, isomalt,maltose, tagatose, lactose, inulin, glycerol, propylene glycol, polyols,polydextrose, fructooligosaccharides, cellulose and cellulosederivatives, and the like, and mixtures thereof. Additionally, inaccordance with still other embodiments of the invention, granulatedsugar (sucrose) or other caloric sweeteners such as crystallinefructose, other carbohydrates, or sugar alcohol can be used as a bulkingagent due to their provision of good content uniformity without theaddition of significant calories.

As used herein, the phrase “anti-caking agent” and “flow agent” refer toany composition which assists in content uniformity and uniformdissolution. In accordance with particular embodiments, non-limitingexamples of anti-caking agents include cream of tartar, calciumsilicate, silicon dioxide, microcrystalline cellulose (Avicel, FMCBioPolymer, Philadelphia, Pa.), and tricalcium phosphate. In oneembodiment, the anti-caking agents are present in the tabletopfunctional sweetener composition in an amount from about 0.001 to about3% by weight of the tabletop functional sweetener composition.

The tabletop sweetener compositions can be packaged in any form known inthe art. Non-limiting forms include, but are not limited to, powderform, granular form, packets, tablets, sachets, pellets, cubes, solids,and liquids.

In one embodiment, the tabletop sweetener composition is asingle-serving (portion control) packet comprising a dry-blend.Dry-blend formulations generally may comprise powder or granules.Although the tabletop sweetener composition may be in a packet of anysize, an illustrative non-limiting example of conventional portioncontrol tabletop sweetener packets are approximately 2.5 by 1.5 inchesand hold approximately 1 gram of a sweetener composition having asweetness equivalent to 2 teaspoons of granulated sugar (˜8 g). Theamount of Reb X in a dry-blend tabletop sweetener formulation can vary.In a particular embodiment, a dry-blend tabletop sweetener formulationmay contain Reb X in an amount from about 1% (w/w) to about 10% (w/w) ofthe tabletop sweetener composition.

Solid tabletop sweetener embodiments include cubes and tablets. Anon-limiting example of conventional cubes are equivalent in size to astandard cube of granulated sugar, which is approximately 2.2×2.2×2.2cm³ and weigh approximately 8 g. In one embodiment, a solid tabletopsweetener is in the form of a tablet or any other form known to thoseskilled in the art.

A tabletop sweetener composition also may be embodied in the form of aliquid, wherein Reb X is combined with a liquid carrier. Suitablenon-limiting examples of carrier agents for liquid tabletop functionalsweeteners include water, alcohol, polyol, glycerin base or citric acidbase dissolved in water, and mixtures thereof. The sweetness equivalentof a tabletop sweetener composition for any of the forms describedherein or known in the art may be varied to obtain a desired sweetnessprofile. For example, a tabletop sweetener composition may comprise asweetness comparable to that of an equivalent amount of standard sugar.In another embodiment, the tabletop sweetener composition may comprise asweetness of up to 100 times that of an equivalent amount of sugar. Inanother embodiment, the tabletop sweetener composition may comprise asweetness of up to 90 times, 80 times, 70 times, 60 times, 50 times, 40times, 30 times, 20 times, 10 times, 9 times, 8 times, 7 times, 6 times,5 times, 4 times, 3 times, and 2 times that of an equivalent amount ofsugar.

Beverage and Beverage Products

In one embodiment, the sweetened composition is a beverage product. Asused herein a “beverage product” is a ready-to-drink beverage, abeverage concentrate, a beverage syrup, or a powdered beverage. Suitableready-to-drink beverages include carbonated and non-carbonatedbeverages. Carbonated beverages include, but are not limited to,enhanced sparkling beverages, cola, lemon-lime flavored sparklingbeverage, orange flavored sparkling beverage, grape flavored sparklingbeverage, strawberry flavored sparkling beverage, pineapple flavoredsparkling beverage, ginger-ale, soft drinks and root beer.Non-carbonated beverages include, but are not limited to fruit juice,fruit-flavored juice, juice drinks, nectars, vegetable juice,vegetable-flavored juice, sports drinks, energy drinks, enhanced waterdrinks, enhanced water with vitamins, near water drinks (e.g., waterwith natural or synthetic flavorants), coconut water, tea type drinks(e.g. black tea, green tea, red tea, oolong tea), coffee, cocoa drink,beverage containing milk components (e.g. milk beverages, coffeecontaining milk components, café au lait, milk tea, fruit milkbeverages), beverages containing cereal extracts, smoothies andcombinations thereof.

Beverage concentrates and beverage syrups are prepared with an initialvolume of liquid matrix (e.g. water) and the desired beverageingredients. Full strength beverages are then prepared by adding furthervolumes of water. Powdered beverages are prepared by dry-mixing all ofthe beverage ingredients in the absence of a liquid matrix. Fullstrength beverages are then prepared by adding the full volume of water.

Beverages comprise a liquid matrix, i.e. the basic ingredient in whichthe ingredients—including the sweetener or sweetener compositions—aredissolved. In one embodiment, a beverage comprises water of beveragequality as the liquid matrix, such as, for example deionized water,distilled water, reverse osmosis water, carbon-treated water, purifiedwater, demineralized water and combinations thereof, can be used.Additional suitable liquid matrices include, but are not limited tophosphoric acid, phosphate buffer, citric acid, citrate buffer andcarbon-treated water.

In one embodiment, a beverage contains Reb X as the sole sweetener.

In another embodiment, a beverage contains a sweetener compositioncomprising Reb X. Any sweetener composition comprising Reb X detailedherein can be used in the beverages.

In another embodiment, a method of preparing a beverage comprisescombining a liquid matrix and Reb X. The method can further compriseaddition of one or more sweeteners, additives and/or functionalingredients.

In still another embodiment, a method of preparing a beverage comprisescombining a liquid matrix and a sweetener composition comprising Reb X.

In one embodiment, a beverage contains Reb X in an amount ranging fromabout 1 ppm to about 10,000 ppm, such as, for example, from about 25 ppmto about 800 ppm. In another embodiment, Reb X is present in a beveragein an amount ranging from about 100 ppm to about 600 ppm. In yet otherembodiments, Reb X is present in a beverage in an amount ranging fromabout 100 to about 200 ppm, from about 100 ppm to about 300 ppm, fromabout 100 ppm to about 400 ppm, or from about 100 ppm to about 500 ppm.In still another embodiment, Reb X is present in a beverage in an amountranging from about 300 to about 700 ppm, such as, for example, fromabout 400 ppm to about 600 ppm. In a particular embodiment, Reb X ispresent in a beverage an amount of about 500 ppm.

In another embodiment, a beverage contains a sweetener compositioncontaining Reb X, wherein Reb X is present in the beverage in an amountranging from about 1 ppm to about 10,000 ppm, such as, for example, fromabout 25 ppm to about 800 ppm. In another embodiment, Reb X is presentin the beverage in an amount ranging from about 100 ppm to about 600ppm. In yet other embodiments, Reb X is present in the beverage in anamount ranging from about 100 to about 200 ppm, from about 100 ppm toabout 300 ppm, from about 100 ppm to about 400 ppm, or from about 100ppm to about 500 ppm. In still another embodiment, Reb X is present inthe beverage in an amount ranging from about 300 to about 700 ppm, suchas, for example, from about 400 ppm to about 600 ppm. In a particularembodiment, Reb X is present in the beverage in an amount of about 500ppm.

The beverage can further include at least one additional sweetener. Anyof the sweeteners detailed herein can be used, including natural,non-natural, or synthetic sweeteners.

In one embodiment, carbohydrate sweeteners can be present in thebeverage in a concentration from about 100 ppm to about 140,000 ppm.Synthetic sweeteners may be present in the beverage in a concentrationfrom about 0.3 ppm to about 3,500 ppm. Natural high potency sweetenersmay be present in the beverage in a concentration from about 0.1 ppm toabout 3,000 ppm.

The beverage can further include additives including, but are notlimited to, carbohydrates, polyols, amino acids and their correspondingsalts, poly-amino acids and their corresponding salts, sugar acids andtheir corresponding salts, nucleotides, organic acids, inorganic acids,organic salts including organic acid salts and organic base salts,inorganic salts, bitter compounds, caffeine, flavorants and flavoringingredients, astringent compounds, proteins or protein hydrolysates,surfactants, emulsifiers, weighing agents, juice, dairy, cereal andother plant extracts, flavonoids, alcohols, polymers and combinationsthereof. Any suitable additive described herein can be used.

In one embodiment, the polyol can be present in the beverage in aconcentration from about 100 ppm to about 250,000 ppm, such as, forexample, from about 5,000 ppm to about 40,000 ppm.

In another embodiment, the amino acid can be present in the beverage ina concentration from about 10 ppm to about 50,000 ppm, such as, forexample, from about 1,000 ppm to about 10,000 ppm, from about 2,500 ppmto about 5,000 ppm or from about 250 ppm to about 7,500 ppm.

In still another embodiment, the nucleotide can be present in thebeverage in a concentration from about 5 ppm to about 1,000 ppm.

In yet another embodiment, the organic acid additive can be present inthe beverage in a concentration from about 10 ppm to about 5,000 ppm.

In yet another embodiment, the inorganic acid additive can be present inthe beverage in a concentration from about 25 ppm to about 25,000 ppm.

In still another embodiment, the bitter compound can be present in thebeverage in a concentration from about 25 ppm to about 25,000 ppm.

In yet another embodiment, the flavorant can be present in the beveragea concentration from about 0.1 ppm to about 4,000 ppm.

In a still further embodiment, the polymer can be present in thebeverage in a concentration from about 30 ppm to about 2,000 ppm.

In another embodiment, the protein hydrosylate can be present in thebeverage in a concentration from about 200 ppm to about 50,000.

In yet another embodiment, the surfactant additive can be present in thebeverage in a concentration from about 30 ppm to about 2,000 ppm.

In still another embodiment, the flavonoid additive can be present inthe beverage a concentration from about 0.1 ppm to about 1,000 ppm.

In yet another embodiment, the alcohol additive can be present in thebeverage in a concentration from about 625 ppm to about 10,000 ppm.

In a still further embodiment, the astringent additive can be present inthe beverage in a concentration from about 10 ppm to about 5,000 ppm.

The beverage can further contain one or more functional ingredients,detailed above. Functional ingredients include, but are not limited to,vitamins, minerals, antioxidants, preservatives, glucosamine,polyphenols and combinations thereof. Any suitable functional ingredientdescribed herein can be used.

It is contemplated that the pH of the sweetened composition, such as,for example, a beverage, does not materially or adversely affect thetaste of the sweetener. A non-limiting example of the pH range of thesweetenable composition may be from about 1.8 to about 10. A furtherexample includes a pH range from about 2 to about 5. In a particularembodiment, the pH of beverage can be from about 2.5 to about 4.2. On ofskill in the art will understand that the pH of the beverage can varybased on the type of beverage. Dairy beverages, for example, can havepHs greater than 4.2.

The titratable acidity of a beverage comprising Reb X may, for example,range from about 0.01 to about 1.0% by weight of beverage.

In one embodiment, the sparkling beverage product has an acidity fromabout 0.01 to about 1.0% by weight of the beverage, such as, forexample, from about 0.05% to about 0.25% by weight of beverage.

The carbonation of a sparkling beverage product has 0 to about 2% (w/w)of carbon dioxide or its equivalent, for example, from about 0.1 toabout 1.0% (w/w).

The temperature of a beverage comprising Reb X may, for example, rangefrom about 4° C. to about 100° C., such as, for example, from about 4°C. to about 25° C.

The beverage can be a full-calorie beverage that has up to about 120calories per 8 oz serving.

The beverage can be a mid-calorie beverage that has up to about 60calories per 8 oz serving.

The beverage can be a low-calorie beverage that has up to about 40calories per 8 oz serving.

The beverage can be a zero-calorie that has less than about 5 caloriesper 8 oz. serving.

In one embodiment, a beverage comprises between about 200 ppm and about500 ppm Reb X, wherein the liquid matrix of the beverage is selectedfrom the group consisting of water, acidified water, phosphoric acid,phosphate buffer, citric acid, citrate buffer, carbon-treated water andcombinations thereof. The pH of the beverage can be from about 2.5 toabout 4.2. The beverage can further include additives, such as, forexample, erythritol. The beverage can further include functionalingredients, such as, for example vitamins.

In particular embodiments, a beverage comprises Reb X; a polyol selectedfrom erythritol, maltitol, mannitol, xylitol, glycerol, sorbitol, andcombinations thereof; and optionally at least one additional sweetenerand/or functional ingredient. In a particular embodiment, the polyol iserythritol. In one embodiment, Reb X and the polyol are present in thebeverage in a weight ratio from about 1:1 to about 1:800, such as, forexample, from about 1:4 to about 1:800, from about 1:20 to about 1:600,from about 1:50 to about 1:300 or from about 1:75 to about 1:150. Inanother embodiment, Reb X is present in the beverage in a concentrationfrom about 1 ppm to about 10,000 ppm, such as, for example, about 500ppm. The polyol, such as, for example, erythritol, is present in thebeverage in a concentration from about 100 ppm to about 250,000 ppm,such as, for example, from about 5,000 ppm to about 40,000 ppm, fromabout 1,000 ppm to about 35,000 ppm.

In a particular embodiment, a beverage comprises a sweetener compositioncomprising Reb X and erythritol as the sweetener component of thesweetener composition. Generally, erythritol can comprise from about0.1% to about 3.5% by weight of the sweetener component. Reb X can bepresent in the beverage in a concentration from about 50 ppm to about600 ppm and erythritol can be from about 0.1% to about 3.5% by weight ofthe sweetener component. In a particular embodiment, the concentrationof Reb X in the beverage is about 300 ppm and erythritol is 0.1% toabout 3.5% by weight of the sweetener component. The pH of the beverageis preferably between about 2.5 to about 4.2.

In particular embodiments, a beverage comprises Reb X; a carbohydratesweetener selected from sucrose, fructose, glucose, maltose andcombinations thereof; and optionally at least one additional sweetenerand/or functional ingredient. The Reb X can be provided as a purecompound or as part of a Stevia extract or steviol glycoside mixture, asdescribed above. Reb X can be present in an amount from about 5% toabout 99% by weight on a dry basis in either a steviol glycoside mixtureor a Stevia extract. In one embodiment, Reb X and the carbohydrate arepresent in a sweetener composition in a weight ratio from about 0.001:14to about 1:0.01, such as, for example, about 0.06:6. In one embodiment,Reb X is present in the beverage in a concentration from about 1 ppm toabout 10,000 ppm, such as, for example, about 500 ppm. The carbohydrate,such as, for example, sucrose, is present in the beverage aconcentration from about 100 ppm to about 140,000 ppm, such as, forexample, from about 1,000 ppm to about 100,000 ppm, from about 5,000 ppmto about 80,000 ppm.

In particular embodiments, a beverage comprises Reb X; an amino acidselected from glycine, alanine, proline, taurine and combinationsthereof; and optionally at least one additional sweetener and/orfunctional ingredient. In one embodiment, Reb X is present in thebeverage in a concentration from about 1 ppm to about 10,000 ppm, suchas, for example, about 500 ppm. The amino acid, such as, for example,glycine, can be present in the beverage in a concentration from about 10ppm to about 50,000 ppm when present in a sweetened composition, suchas, for example, from about 1,000 ppm to about 10,000 ppm, from about2,500 ppm to about 5,000 ppm

In particular embodiments, a beverage comprises Reb X; a salt selectedfrom sodium chloride, magnesium chloride, potassium chloride, calciumchloride, phosphate salts and combinations thereof; and optionally atleast one additional sweetener and/or functional ingredient. In oneembodiment, Reb X is present in the beverage in a concentration fromabout 1 ppm to about 10,000 ppm, such as, for example, about 500 ppm.The inorganic salt, such as, for example, magnesium chloride, is presentin the beverage in a concentration from about 25 ppm to about 25,000ppm, such as, for example, from about 100 ppm to about 4,000 ppm or fromabout 100 ppm to about 3,000 ppm.

In another embodiment, a beverage comprises a sweetener compositioncomprising Reb X and Reb B as the sweetener component of the sweetenercomposition. The relative weight percent of Reb X and Reb B can eachvary from about 1% to about 99% when dry, such as for example, about 95%Reb X/5% Reb B, about 90% Reb X/10% Reb B, about 85% Reb X/15% Reb B,about 80% Reb X/20% Reb B, about 75% Reb X/25% Reb B, about 70% RebX/30% Reb B, about 65% Reb X/35% Reb B, about 60% Reb X/40% Reb B, about55% Reb X/45% Reb B, about 50% Reb X/50% Reb B, about 45% Reb X/55% RebB, about 40% Reb X/60% Reb B, about 35% Reb X/65% Reb B, about 30% RebX/70% Reb B, about 25% Reb X/75% Reb B, about 20% Reb X/80% Reb B, about15% Reb X/85% Reb B, about 10% Reb X/90% Reb B or about 5% Reb X/10% RebB. In a particular embodiment, Reb B comprises from about 5% to about40% by weight of the sweetener component, such as, for example, fromabout 10% to about 30% or about 15% to about 25%. In another particularembodiment, Reb X is present in the beverage in a concentration fromabout 50 ppm to about 600 ppm, such as, for example, from about 100 toabout 400 ppm, and Reb B comprises from about 5% to about 40% by weightof the sweetener component. In another embodiment, Reb X is present in aconcentration from about 50 ppm to about 600 ppm and Reb B is present ina concentration from about 10 to about 150 ppm. In a more particularembodiment, Reb X is present in a concentration of about 300 ppm and RebB is present in a concentration from about 50 ppm to about 100 ppm. ThepH of the beverage is preferably between about 2.5 to about 4.2.

In another embodiment, a beverage comprises a sweetener compositioncomprises Reb X and NSF-02 (available from PureCircle) as the sweetenercomponent of the sweetener composition. The relative weight percent ofReb X and NSF-02 can each vary from about 1% to about 99%, such as forexample, about 95% Reb X/5% NSF-02, about 90% Reb X/10% NSF-02, about85% Reb X/15% NSF-02, about 80% Reb X/20% NSF-02, about 75% Reb X/25%NSF-02, about 70% Reb X/30% NSF-02, about 65% Reb X/35% NSF-02, about60% Reb X/40% NSF-02, about 55% Reb X/45% NSF-02, about 50% Reb X/50%NSF-02, about 45% Reb X/55% NSF-02, about 40% Reb X/60% NSF-02, about35% Reb X/65% NSF-02, about 30% Reb X/70% NSF-02, about 25% Reb X/75%NSF-02, about 20% Reb X/80% NSF-02, about 15% Reb X/85% NSF-02, about10% Reb X/90% NSF-02 or about 5% Reb X/10% NSF-02. In a particularembodiment, NSF-02 comprises from about 5% to about 50% by weight of thesweetener component, such as, for example, from about 10% to about 40%or about 20% to about 30%. In another particular embodiment, Reb X ispresent in the beverage in a concentration from about 50 ppm to about600 ppm, such as, for example, from about 100 to about 400 ppm, andNSF-02 comprises from about 5% to about 50% by weight of the sweetenercomponent. In a more particular embodiment, Reb X is present in aconcentration from about 50 ppm to about 600 ppm and NSF-02 is presentin a concentration from about 10 ppm about 150 ppm. In a more particularembodiment, Reb X is present in a concentration of about 300 ppm andNSF-02 is present in a concentration from about 25 ppm to about 100 ppm.The pH of the beverage is preferably between about 2.5 to about 4.2.

In still another embodiment, a beverage comprises a sweetenercomposition comprises Reb X and mogroside V as the sweetener componentof the sweetener composition. The relative weight percent of Reb X andmogroside V can each vary from about 1% to about 99%, such as forexample, about 95% Reb X/5% mogroside V, about 90% Reb X/10% mogrosideV, about 85% Reb X/15% mogroside V, about 80% Reb X/20% mogroside V,about 75% Reb X/25% mogroside V, about 70% Reb X/30% mogroside V, about65% Reb X/35% mogroside V, about 60% Reb X/40% mogroside V, about 55%Reb X/45% mogroside V, about 50% Reb X/50% mogroside V, about 45% RobX/55% mogroside V, about 40% Reb X/60% mogroside V, about 35% Reb X/65%mogroside V, about 30% Rob X/70% mogroside V, about 25% Reb X/75%mogroside V, about 20% Reb X/80% mogroside V, about 15% Reb X/85%mogroside V, about 10% Reb X/90% mogroside V or about 5% Reb X/10%mogroside V. In a particular embodiment, mogroside V comprises fromabout 5% to about 50% of the sweetener component, such as, for example,from about 10% to about 40% or about 20% to about 30%. In anotherparticular embodiment, Reb X is present in the beverage in aconcentration from about 50 ppm to about 600 ppm, such as, for example,from about 100 to about 400 ppm, and mogroside V comprises from about 5%to about 50% by weight of the sweetener component. In a more particularembodiment, Reb X is present in a concentration from about 50 ppm toabout 600 ppm and mogroside V is present in a concentration from about10 ppm about 250 ppm. In a more particular embodiment, Reb X is presentin a concentration of about 300 ppm and mogroside is present in aconcentration from about 100 ppm to about 200 ppm. The pH of thebeverage is preferably between about 2.5 to about 4.2.

In another embodiment, a beverage comprises a sweetener compositioncomprises Reb X and Reb A as the sweetener component of the sweetenercomposition. The relative weight percent of Reb X and Reb A can eachvary from about 1% to about 99%, such as for example, about 95% Reb X/5%Reb A, about 90% Reb X/10% Reb A, about 85% Reb X/15% Reb A, about 80%Reb X/20% Reb A, about 75% Reb X/25% Reb A, about 70% Reb X/30% Reb A,about 65% Reb X/35% Reb A, about 60% Reb X/40% Reb A, about 55% RebX/45% Reb A, about 50% Reb X/50% Reb A, about 45% Reb X/55% Reb A, about40% Reb X/60% Reb A, about 35% Reb X/65% Reb A, about 30% Reb X/70% RebA, about 25% Reb X/75% Reb A, about 20% Reb X/80% Reb A, about 15% RobX/85% Reb A, about 10% Rob X/90% Rob A or about 5% Reb X/10% Reb A. In aparticular embodiment, Reb A comprises from about 5% to about 40% of thesweetener component, such as, for example, from about 10% to about 30%or about 15% to about 25%. In another particular embodiment, Reb X ispresent in the beverage in a concentration from about 50 ppm to about600 ppm, such as, for example, from about 100 to about 400 ppm, and RebA comprises from about 5% to about 40% by weight of the sweetenercomponent. In another embodiment, Rob X is present in a concentrationfrom about 50 ppm to about 600 ppm and Reb A is present in aconcentration from about 10 to about 500 ppm. In a more particularembodiment, Rob X is present in a concentration of about 300 ppm and RebA is present in a concentration from of about 100 ppm. The pH of thebeverage is preferably between about 2.5 to about 4.2.

In another embodiment, a beverage comprises a sweetener compositioncomprising Reb X and Reb D as the sweetener component of the sweetenercomposition. The relative weight percent of Reb X and Reb D can eachvary from about 1% to about 99%, such as for example, about 95% Reb X/5%Reb D, about 90% Reb X/10% Reb D, about 85% Reb X/15% Reb D, about 80%Reb X/20% Reb D, about 75% Reb X/25% Reb D, about 70% Reb X/30% Reb D,about 65% Reb X/35% Reb D, about 60% Reb X/40% Reb D, about 55% RebX/45% Reb D, about 50% Reb X/50% Reb D, about 45% Reb X/55% Reb D, about40% Reb X/60% Reb D, about 35% Reb X/65% Reb D, about 30% Reb X/70% RobD, about 25% Reb X/75% Reb D, about 20% Reb X/80% Reb D, about 15% RebX/85% Reb D, about 10% Rob X/90% Reb D or about 5% Reb X/10% Reb D. In aparticular embodiment, Reb D comprises from about 5% to about 40% of thesweetener component, such as, for example, from about 10% to about 30%or about 15% to about 25%. In another particular embodiment, Reb X ispresent in the beverage in a concentration from about 50 ppm to about600 ppm, such as, for example, from about 100 to about 400 ppm, and RebD comprises from about 5% to about 40% by weight of the sweetenercomponent. In another embodiment, Reb X is present in a concentrationfrom about 50 ppm to about 600 ppm and Reb D is present in aconcentration from about 10 ppm to about 500 ppm. In a more particularembodiment, Reb X is present in a concentration of about 300 ppm and RebD is present in a concentration from of about 100 ppm. The pH of thebeverage is preferably between about 2.5 to about 4.2.

In another embodiment, a beverage comprises a sweetener compositioncomprises Reb X, Reb A and Reb D as the sweetener component of thesweetener composition. The relative weight percent of Reb X, Reb A andReb D can each vary from about 1% to about 99%. In a particularembodiment, Reb A and Reb D together comprise from about 5% to about 40%of the sweetener component, such as, for example, from about 10% toabout 30% or about 15% to about 25%. In another particular embodiment,Reb X is present in the beverage in a concentration from about 50 ppm toabout 600 ppm, such as, for example, from about 100 to about 400 ppm,and Reb A and Reb D together comprise from about 5% to about 40% byweight of the sweetener component. In another embodiment, Reb X ispresent in a concentration from about 50 ppm to about 600 ppm, Reb A ispresent in a concentration from about 10 ppm to about 500 ppm and Reb Dis present in a concentration from about 10 ppm to about 500 ppm. In amore particular embodiment, Reb X is present in a concentration of about200 ppm, Reb A is present in a concentration of about 100 ppm and Reb Dis present in a concentration from of about 100 ppm. The pH of thebeverage is preferably between about 2.5 to about 4.2.

In another embodiment, a beverage comprises a sweetener compositioncomprises Reb X, Reb B and Reb D as the sweetener component of thesweetener composition. The relative weight percent of Reb X, Reb B andReb D can each vary from about 1% to about 99%. In a particularembodiment, Reb B and Reb D together comprise from about 5% to about 40%of the sweetener component, such as, for example, from about 10% toabout 30% or about 15% to about 25%. In another particular embodiment,Reb X is present in the beverage in a concentration from about 50 ppm toabout 600 ppm, such as, for example, from about 100 to about 400 ppm,and Reb B and Reb D together comprise from about 5% to about 40% byweight of the sweetener component. In another embodiment, Reb X ispresent in a concentration from about 50 ppm to about 600 ppm, Reb B ispresent in a concentration from about 10 ppm to about 500 ppm and Reb Dis present in a concentration from about 10 ppm to about 500 ppm. In amore particular embodiment, Reb X is present in a concentration of about200 ppm, Reb B is present in a concentration of about 100 ppm and Reb Dis present in a concentration from of about 100 ppm. The pH of thebeverage is preferably between about 2.5 to about 4.2.

Methods for Improving Temporal and/or Flavor Profile

A method for imparting a more sugar-like temporal profile, flavorprofile, or both to a sweetenable composition comprises combining asweetenable composition with Reb X or the sweetener compositions of thepresent invention, i.e., sweetener compositions containing Reb X.

The method can further include the addition of other sweeteners,additives, functional ingredients and combinations thereof. Anysweetener, additive or functional ingredient detailed herein can beused.

As used herein, the “sugar-like” characteristics include anycharacteristic similar to that of sucrose and include, but are notlimited to, maximal response, flavor profile, temporal profile,adaptation behavior, mouthfeel, concentration/response function,tastant/and flavor/sweet taste interactions, spatial patternselectivity, and temperature effects.

The flavor profile of a sweetener is a quantitative profile of therelative intensities of all of the taste attributes exhibited. Suchprofiles often are plotted as histograms or radar plots.

These characteristics are dimensions in which the taste of sucrose isdifferent from the tastes of Reb X. Of these, however, the flavorprofile and temporal profile are particularly important. In a singletasting of a sweet food or beverage, differences (1) in the attributesthat constitute a sweetener's flavor profile and (2) in the rates ofsweetness onset and dissipation, which constitute a sweetener's temporalprofile, between those observed for sucrose and for Reb X can be noted.

Whether or not a characteristic is more sugar-like is determined by anexpert sensory panel who taste compositions comprising sugar andcompositions comprising Reb X, both with and without additives, andprovide their impression as to the similarities of the characteristicsof the sweetener compositions, both with and without additives, withthose comprising sugar. A suitable procedure for determining whether acomposition has a more sugar-like taste is described in embodimentsdescribed herein below.

In a particular embodiment, a panel of assessors is used to measure thereduction of sweetness linger. Briefly described, a panel of assessors(generally 8 to 12 individuals) is trained to evaluate sweetnessperception and measure sweetness at several time points from when thesample is initially taken into the mouth until 3 minutes after it hasbeen expectorated. Using statistical analysis, the results are comparedbetween samples containing additives and samples that do not containadditives. A decrease in score for a time point measured after thesample has cleared the mouth indicates there has been a reduction insweetness perception.

The panel of assessors may be trained using procedures well known tothose of ordinary skill in the art. In a particular embodiment, thepanel of assessors may be trained using the Spectrum™ DescriptiveAnalysis Method (Meilgaard et al, Sensory Evaluation Techniques, 3^(rd)edition, Chapter 11). Desirably, the focus of training should be therecognition of and the measure of the basic tastes; specifically, sweet.In order to ensure accuracy and reproducibility of results, eachassessor should repeat the measure of the reduction of sweetness lingerabout three to about five times per sample, taking at least a fiveminute break between each repetition and/or sample and rinsing well withwater to clear the mouth.

Generally, the method of measuring sweetness comprises taking a 10 mLsample into the mouth, holding the sample in the mouth for 5 seconds andgently swirling the sample in the mouth, rating the sweetness intensityperceived at 5 seconds, expectorating the sample (without swallowingfollowing expectorating the sample), rinsing with one mouthful of water(e.g., vigorously moving water in mouth as if with mouth wash) andexpectorating the rinse water, rating the sweetness intensity perceivedimmediately upon expectorating the rinse water, waiting 45 seconds and,while waiting those 45 seconds, identifying the time of maximumperceived sweetness intensity and rating the sweetness intensity at thattime (moving the mouth normally and swallowing as needed), rating thesweetness intensity after another 10 seconds, rating the sweetnessintensity after another 60 seconds (cumulative 120 seconds after rinse),and rating the sweetness intensity after still another 60 seconds(cumulative 180 seconds after rinse). Between samples take a 5 minutebreak, rinsing well with water to clear the mouth.

Delivery Systems

Reb X and sweetener compositions comprising Reb X can also be formulatedinto various delivery systems having improved ease of handling and rateof dissolution. Non-limiting examples of suitable delivery systemscomprise sweetener compositions co-crystallized with a sugar or apolyol, agglomerated sweetener compositions, compacted sweetenercompositions, dried sweetener compositions, particle sweetenercompositions, spheronized sweetener compositions, granular sweetenercompositions, and liquid sweetener compositions.

Co-Crystallized Sugar/Polyol and Reb X Composition

In a particular embodiment, a sweetener composition is co-crystallizedwith a sugar or a polyol in various ratios to prepare a substantiallywater soluble sweetener with substantially no dusting problems. Sugar,as used herein, generally refers to sucrose (C₁₂H₂₂O₁₁). Polyol, as usedherein, is synonymous with sugar alcohol and generally refers to amolecule that contains more than one hydroxyl group, erythritol,maltitol, mannitol, sorbitol, lactitol, xylitol, isomalt, propyleneglycol, glycerol (glycerine), threitol, galactitol, palatinose, reduceisomalto-oligosaccharides, reduced xylo-oligosaccharides, reducedgentio-oligosaccharides, reduced maltose syrup, reduced glucose syrup,and sugar alcohols or any other carbohydrates capable of being reducedwhich do not adversely affect the taste of the sweetener composition.

In another embodiment, a process for preparing a sugar or a polyolco-crystallized Reb X sweetener composition is provided. Such methodsare known to those of ordinary skill in the art, and are discussed inmore detail in U.S. Pat. No. 6,214,402. According to certainembodiments, the process for preparing a sugar or a polyolco-crystallized Reb X sweetener composition may comprise the steps ofpreparing a supersaturated sugar or polyol syrup, adding a predeterminedamount of premix comprising a desired ratio of the Reb X sweetenercomposition and sugar or polyol to the syrup with vigorous mechanicalagitation, removing the sugar or polyol syrup mixture from heat, andquickly cooling the sugar or polyol syrup mixture with vigorousagitation during crystallization and agglomeration. During the processthe Reb X sweetener composition is incorporated as an integral part ofthe sugar or polyol matrix, thereby preventing the sweetener compositionfrom separating or settling out of the mixture during handling,packaging, or storing. The resulting product may be granular,free-flowing, non-caking, and may be readily and uniformly dispersed ordissolved in water.

In a particular embodiment, a sugar or a polyol syrup may be obtainedcommercially or by effectively mixing a sugar or a polyol with water.The sugar or polyol syrup may be supersaturated to produce a syrup witha solids content in the range of about 95 to about 98% by weight of thesyrup by removing water from the sugar syrup. Generally, the water maybe removed from the sugar or polyol syrup by heating and agitating thesugar or polyol syrup while maintaining the sugar or polyol syrup at atemperature of not less than about 120° C. to prevent prematurecrystallization.

In another particular embodiment, a dry premix is prepared by combiningthe Reb X sweetener composition and a sugar or a polyol in a desiredamount. According to certain embodiments, the weight ratio of the Reb Xsweetener composition to sugar or polyol is in the range of about0.001:1 to about 1:1. Other components, such as flavors or otherhigh-potency sweeteners, also may be added to the dry premix, so long asthe amount does not adversely affect the overall taste of the sugarco-crystallized sweetener composition.

The amounts of premix and supersaturated syrup may be varied in order toproduce products with varying levels of sweetness. In particularembodiments, the Reb X sweetener composition is present in an amountfrom about 0.001% to about 50% by weight of the final product, or fromabout 0.001% to about 5%, or from about 0.001% to about 2.5%.

The sugar or polyol co-crystallized sweetener compositions of thisinvention are suitable for use in any sweetenable composition to replaceconventional caloric sweeteners, as well as other types of low-caloricor non-caloric sweeteners. In addition, the sugar or polyolco-crystallized sweetener composition described herein can be combinedin certain embodiments with bulking agents, non-limiting examples ofwhich include dextrose, maltodextrin, lactose, inulin, polyols,polydextrose, cellulose and cellulose derivatives. Such products may beparticularly suitable for use as tabletop sweeteners.

Agglomerated Sweetener Composition

In certain embodiments, an agglomerate of a Reb X sweetener compositionis provided. As used herein, “sweetener agglomerate” means a pluralityof sweetener particles clustered and held together. Examples ofsweetener agglomerates include, but are not limited to, binder heldagglomerates, extrudates, and granules.

Binder Held Agglomerates

According to certain embodiments, a process for preparing an agglomerateof a Reb X sweetener composition, a binding agent and a carrier isprovided. Methods for making agglomerates are known to those of ordinaryskill in the art, and are disclosed in more detail in U.S. Pat. No.6,180,157. Generally described, the process for preparing an agglomeratein accordance with a certain embodiment comprises the steps of preparinga premix solution comprising a Reb X sweetener composition and a bindingagent in a solvent, heating the premix to a temperature sufficient toeffectively form a mixture of the premix, applying the premix onto afluidized carrier by a fluid bed agglomerator, and drying the resultingagglomerate. The sweetness level of the resulting agglomerate may bemodified by varying the amount of the sweetener composition in thepremix solution.

In a particular embodiment, the premix solution comprises a Reb Xsweetener composition and a binding agent dissolved in a solvent. Thebinding agent may have sufficient binding strength to facilitateagglomeration. Non-limiting examples of suitable binding agents includemaltodextrin, sucrose, gellan gum, gum arabic, hydroxypropylmethylcellulose, carboxymethyl cellulose, polyvinyl pyrrolidone, cellobiose,proteins and mixtures thereof. The Reb X sweetener composition andbinding agent may be dissolved in the same solvent or in two separatesolvents. In embodiments wherein separate solvents are used to dissolvethe sweetener composition and binding agent, the solvents may be thesame or different before being combined into a single solution. Anysolvent in which the Reb X sweetener composition and/or binding agentdissolves may be used. Desirably, the solvent is a food grade solvent,non-limiting examples of which include ethanol, water, isopropanol,methanol, and mixtures thereof. In order to effect complete mixing ofthe premix, the premix may be heated up to a temperature in the range ofabout 30 to about 100° C. As used herein, the term “effect mixing” meansblending sufficiently so as to form a mixture.

The amount of binding agent in the solution may vary depending on avariety of factors, including the binding strength of the particularbinding agent and the particular solvent chosen. The binding agent isgenerally present in the premix solution in an amount from about 1 toabout 50% by weight of the premix solution, or from about 5 to about 25%by weight. The weight ratio of the binding agent to the Reb X sweetenercomposition in the premix solution may vary from as low as about 1:10 toas high as about 10:1. The weight ratio of the binding agent to the RebX sweetener composition may also vary from about 0.5:1.0 to about 2:1.

Following preparation of the premix solution, the premix solution isapplied onto a fluidized carrier using a fluid bed agglomeration mixer.Preferably, the premix is applied onto the fluidized carrier by sprayingthe premix onto the fluidized carrier to form an agglomerate of the RebX sweetener composition and the carrier. The fluid bed agglomerator maybe any suitable fluid bed agglomerator known to those of ordinary skillin the art. For example, the fluid bed agglomerator may be a batch, acontinuous, or a continuous turbulent flow agglomerator.

The carrier is fluidized and its temperature is adjusted to betweenabout 20 and about 50° C., or to between about 35 and about 45° C. In acertain embodiments, the carrier is heated to about 40° C. The carriermay be placed into a removable bowl of a fluid bed agglomerator. Afterthe bowl is secured to the fluid bed agglomerator, the carrier isfluidized and heated as necessary by adjusting the inlet airtemperature. The temperature of the inlet air can be maintained betweenabout 50 and about 100° C. For example, to heat the fluidized carrier toabout 40° C., the inlet air temperature may be adjusted to between about70 and about 75° C.

Once the fluidized carrier reaches the desired temperature, the premixsolution may be applied through the spray nozzle of the fluid bedagglomerator. The premix solution may be sprayed onto the fluidizedcarrier at any rate which is effective to produce an agglomerate havingthe desired particle size distribution. Those skilled in the art willrecognize that a number of parameters may be adjusted to obtain thedesired particle size distribution. After spraying is completed, theagglomerate may be allowed to dry. In certain embodiments, theagglomerate is allowed to dry until the outlet air temperature reachesabout 35 to about 40° C.

The amount of the Reb X sweetener composition, carrier, and bindingagent in the resulting agglomerates may be varied depending on a varietyof factors, including the selection of binding agent and carrier as wellas the desired sweetening potency of the agglomerate. Those of ordinaryskill in the art will appreciate that the amount of Reb X sweetenercomposition present in the agglomerates may be controlled by varying theamount of the Reb X sweetener composition that is added to the premixsolution. The amount of sweetness is particularly important when tryingto match the sweetness delivered by other natural and/or syntheticsweeteners in a variety of products.

In one embodiment, the weight ratio of the carrier to the Reb Xsweetener composition is between about 1:10 and about 10:1, or betweenabout 0.5:1.0 and about 2:1. In one embodiment, the Reb X sweetenercomposition is present in the agglomerates in an amount in the range ofabout 0.1 to about 99.9% by weight, the carrier is present in theagglomerates in an amount in the range of about 50 to about 99.9% byweight, and the amount of binding agent is present in the agglomeratesin an amount in the range of about 0.1 to about 15% by weight based onthe total weight of the agglomerate. In another embodiment, the amountof the Reb X sweetener composition present in the agglomerate is in therange of about 50 to about 99.9% by weight, the amount of carrierpresent in the agglomerate is in the range of about 75 to about 99% byweight, and the amount of binding agent present in the agglomerate is inthe range of about 1 to about 7% by weight.

The particle size distribution of the agglomerates may be determined bysifting the agglomerate through screens of various sizes. The productalso may be screened to produce a narrower particle size distribution,if desired. For example, a 14 mesh screen may be used to remove largeparticles and produce a product of especially good appearance, particlessmaller than 120 mesh may be removed to obtain an agglomerate withimproved flow properties, or a narrower particle size distribution maybe obtained if desired for particular applications.

Those of ordinary skill in the art will appreciate that the particlesize distribution of the agglomerate may be controlled by a variety offactors, including the selection of binding agent, the concentration ofthe binding agent in solution, the spray rate of the spray solution, theatomization air pressure, and the particular carrier used. For example,increasing the spray rate may increase the average particle size.

In certain embodiments, the agglomerates provided herein may be blendedwith blending agents. Blending agents, as used herein, include a broadrange of ingredients commonly used in foods or beverages, including, butnot limited to, those ingredients used as binding agents, carriers,bulking agents, and sweeteners. For example, the agglomerates may beused to prepare tabletop sweeteners or powdered drink mixes by dryblending the agglomerates of this invention with blending agentscommonly used to prepare tabletop sweeteners or powdered drink mixesusing methods well known to those of ordinary skill in the art.

Extrudates

Also provided in embodiments herein are substantially dustless andsubstantially free-flowing extrudates or extruded agglomerates of theReb X sweetener composition. In accordance with certain embodiments,such particles may be formed with or without the use of binders usingextrusion and spheronization processes. “Extrudates” or “extrudedsweetener composition”, as used herein, refers to cylindrical,free-flowing, relatively non-dusty, mechanically strong granules of theReb X sweetener composition. The terms “spheres” or “spheronizedsweetener composition”, as used herein, refer to relatively spherical,smooth, free-flowing, relatively non-dusty, mechanically stronggranules. Although spheres typically have a smoother surface and may bestronger/harder than extrudates, extrudates offer a cost advantage byrequiring less processing. The spheres and extrudates of this inventionmay be processed further, if desired, to form various other particles,such as, for example, by grinding or chopping.

In another embodiment, a process for making extrudates of the Reb Xsweetener composition is provided. Such methods are known to those ofordinary skill in the art and are described in more detail in U.S. Pat.No. 6,365,216. Generally described, the process of making extrudates ofa Reb X sweetener composition comprises the steps of combining the Reb Xsweetener composition, a plasticizer, and optionally a binder to form awet mass; extruding the wet mass to form extrudates; and drying theextrudates to obtain particles of the Reb X sweetener composition.

Non-limiting examples of suitable plasticizers include, but are notlimited to, water, glycerol, and mixtures thereof. In accordance withcertain embodiments, the plasticizer generally is present in the wetmass in an amount from about 4 to about 45% by weight, or from about 15%to about 35% by weight.

Non-limiting examples of suitable binders include, but are not limitedto, polyvinylpyrollidone (PVP), maltodextrins, microcrystallinecellulose, starches, hydroxypropylmethyl cellulose (HPMC),methylcellulose, hydroxypropyl cellulose (HPC), gum arabic, gelatin,xanthan gum, and mixtures thereof. The binder is generally present inthe wet mass in an amount from about 0.01% to about 45% by weight, orfrom about 0.5% to about 10% by weight.

In a particular embodiment, the binder may be dissolved in theplasticizer to form a binder solution that is later added to the Reb Xsweetener composition and other optional ingredients. Use of the bindersolution provides better distribution of the binder through the wetmass.

Other optional ingredients that may be included in the wet mass includecarriers and additives. One of ordinary skill in the art should readilyappreciate that the carriers and additives may comprise any typical foodingredient and also should readily discern the appropriate amount of agiven food ingredient to achieve a desired flavor, taste, orfunctionality.

Methods of extruding the wet mass to form extrudates are well known tothose of ordinary skill in the art. In a particular embodiment, a lowpressure extruder fitted with a die is used to form the extrudates. Theextrudates can be cut into lengths using a cutting device attached tothe discharge end of the extruder to form extrudates that aresubstantially cylindrical in shape and may have the form of noodles orpellets. The shape and size of the extrudates may be varied dependingupon the shape and size of the die openings and the use of the cuttingdevice.

Following the extrusion of the extrudates, the extrudates are driedusing methods well known to those of ordinary skill in the art. In aparticular embodiment, a fluidized bed dryer is used to dry theextrudates.

Optionally, in a particular embodiment, the extrudates are formed intospheres prior to the step of drying. Spheres are formed by charging theextrudates into a marumerizer, which consists of a vertical hollowcylinder (bowl) with a horizontal rotating disc (friction plate)therein. The rotating disc surface can have a variety of textures suitedfor specific purposes. For example, a grid pattern may be used thatcorresponds to the desired particle size. The extrudates are formed intospheres by contact with the rotating disc and by collisions with thewall of the bowl and between particles. During the forming of thespheres, excess moisture may move to the surface or thixotropic behaviormay be exhibited by the extrudates, requiring a slight dusting with asuitable powder to reduce the probability that the particles will sticktogether.

As previously described, the extrudates of the Reb X sweetenercomposition may be formed with or without the use of a′ binder. Theformation of extrudates without the use of a binder is desirable due toits lower cost and improved product quality. In addition, the number ofadditives in the extrudates is reduced. In embodiments wherein theextrudates are formed without the use of a binder, the method of formingparticles further comprises the step of heating the wet mass of the RebX sweetener composition and plasticizer to promote the binding of thewet mass. Desirably, the wet mass is heated to a temperature from about30 to about 90° C., or from about 40 to about 70° C. Methods of heatingthe wet mass, in accordance with certain embodiments, include, but arenot limited to, an oven, a kneader with a heated jacket, or an extruderwith mixing and heating capabilities.

Granules

In one embodiment, granulated forms of a Reb X sweetener composition areprovided. As used herein, the terms “granules,” “granulated forms,” and“granular forms” are synonymous and refer to free-flowing, substantiallynon-dusty, mechanically strong agglomerates of the Reb X sweetenercomposition.

In another embodiment, a process for making granular forms of a Reb Xsweetener composition is provided. Methods of granulation are known tothose of ordinary skill in the art and are described in more detail inthe PCT Publication WO 01/60842. In some embodiments, such methodsinclude, but are not limited to, spray granulation using a wet binderwith or without fluidization, powder compaction, pulverizing, extrusion,and tumble agglomeration. The preferred method of forming granules ispowder compaction due to its simplicity. Also provided herein arecompacted forms of the sweetener Reb X composition.

In one embodiment, the process of forming granules of the Reb Xsweetener composition comprises the steps of compacting the Reb Xsweetener composition to form compacts; breaking up the compacts to formgranules; and optionally screening the granules to obtain granules ofthe Reb X sweetener composition having a desired particle size.

Methods of compacting the Reb X sweetener composition may beaccomplished using any known compacting techniques. Non-limitingexamples of such techniques include roller compaction, tableting,slugging, ram extrusion, plunger pressing, roller briquetting,reciprocating piston processing, die pressing and pelletting. Thecompacts may take any form that may be subjected to subsequent sizereduction, non-limiting examples of which include flakes, chips,briquets, chunks, and pellets. Those of ordinary skill in the art willappreciate that the shape and appearance of the compacts will varydepending upon the shape and surface characteristics of the equipmentused in the compacting step. Accordingly, the compacts may appearsmooth, corrugated, fluted, or pillow-pocketed, or the like. Inaddition, the actual size and characteristics of the compacts willdepend upon the type of equipment and operation parameters employedduring compaction.

In a particularly desirable embodiment, the Reb X sweetener compositionis compacted into flakes or chips using a roller compactor. Aconventional roller compaction apparatus usually includes a hopper forfeeding the sweetener composition to be compacted and a pair ofcounter-rotating rolls, either or both of which are fixed onto theiraxes with one roll optionally slightly moveable. The Reb X sweetenercomposition is fed to the apparatus through the hopper by gravity or aforce-feed screw. The actual size of the resulting compacts will dependupon the width of the roll and scale of the equipment used. In addition,the characteristics of the compacts, such as hardness, density, andthickness will depend on factors such as pressure, roll speed, feedrate, and feed screw amps employed during the compaction process.

In a particular embodiment, the sweetener composition is deaerated priorto the step of compacting, leading to more effective compaction and theformation of stronger compacts and resultant granules. Deaeration may beaccomplished through any known means, non-limiting examples of whichinclude screw feeding, vacuum deaeration, and combinations thereof.

In another particular embodiment, a dry binder is mixed with the Reb Xsweetener composition prior to compaction. The use of a dry binder mayimprove the strength of the granules and aid in their dispersion inliquids. Suitable dry binders include, but are not limited to,pregelatinized corn starch, microcrystalline cellulose, hydrophilicpolymers (e.g., methyl cellulose, hydroxypropylmethyl cellulose,hydroxypropyl cellulose, polyvinylpyrrolidone, alginates, xanthan gum,gellan gum, and gum arabic) and mixtures thereof. In accordance withcertain embodiments, the dry binder generally is present in an amountfrom about 0.1 to about 40% by weight based on the total weight of themixture of the Reb X sweetener composition and dry binder.

Following the step of compacting, the compacts are broken up to formgranules. Any suitable means of breaking up the compacts may be used,including milling. In one particular embodiment, the breaking up of thecompacts is accomplished in a plurality of steps using a variety ofopening sizes for the milling. In some embodiments, the breaking up ofthe compacts is accomplished in two steps: a course breaking step and asubsequent milling step. The step of breaking up the compacts reducesthe number of “overs” in the granulated sweetener composition. As usedherein, “overs” refers to material larger than the largest desiredparticle size.

The breaking up of the compacts generally results in granules of varyingsizes. Accordingly, it may be desirable to screen the granules to obtaingranules having a desired particle size range. Any conventional meansfor screening particles may be used to screen the granules, includingscreeners and sifters. Following screening, the “fines” optionally maybe recycled through the compactor. As used herein, “fines” refers tomaterial smaller than the smallest desired particle size.

Co-Dried Sweetener Composition

Also provided herein are co-dried Reb X sweetener compositionscomprising a Reb X sweetener composition and one or more co-agents.Co-agent, as used herein, includes any ingredient which is desired to beused with and is compatible with the sweetener composition for theproduct being produced. One skilled in the art will appreciate that theco-agents will be selected based on one or more functionalities whichare desirable for use in the product applications for which thesweetener composition will be used. A broad range of ingredients arecompatible with the sweetener compositions, and can be selected for suchfunctional properties. In one embodiment, the one or more co-agentscomprise the at least one additive of the sweetener compositiondescribed herein below. In another embodiment, the one or more co-agentscomprise a bulking agent, flow agent, encapsulating agent, or a mixturethereof.

In another embodiment, a method of co-drying a Reb X sweetenercomposition and one or more co-agents is provided. Such methods areknown to those of ordinary skill in the art and are described in moredetail in PCT Publication WO 02/05660. Any conventional drying equipmentor technique known to those of ordinary skill in the art may be used toco-dry the Reb X sweetener composition and one or more co-agents.Suitable drying processes include, but are not limited to, spray drying,convection drying, vacuum drum drying, freeze drying, pan drying, andhigh speed paddle drying.

In a particularly desirable embodiment, the Reb X sweetener compositionis spray dried. A solution is prepared of the Reb X sweetenercomposition and one or more desired co-agents. Any suitable solvent ormixture of solvents may be used to prepare the solution, depending onthe solubility characteristics of the Reb X sweetener composition andone or more co-agents. In accordance with certain embodiments, suitablesolvents include, but are not limited to, water, ethanol, and mixturesthereof.

In one embodiment, the solution of the Reb X sweetener composition andone or more co-agents may be heated prior to spray drying. Thetemperature can be selected on the basis of the dissolution propertiesof the dry ingredients and the desired viscosity of the spray dryingfeed solution.

In another embodiment, a non-reactive, non-flammable gas (e.g., carbondioxide) may be added to the solution of the Reb X sweetener compositionand one or more co-agents before atomization. The non-reactive,non-flammable gas can be added in an amount effective to lower the bulkdensity of the resulting spray dried product and to produce a productcomprising hollow spheres.

Methods of spray drying are well known to those of ordinary skill in theart. In one embodiment, the solution of the Reb X sweetener compositionand one or more co-agents is fed through a spray dryer at an air inlettemperature in the range of about 150 to about 350° C. Increasing theair inlet temperature at a constant air flow may result in a producthaving reduced bulk density. The air outlet temperature may range fromabout 70 to about 140° C., in accordance with certain embodiments.Decreasing the air outlet temperature may result in a product having ahigh moisture content which allows for ease of agglomeration in a fluidbed dryer to produce sweetener compositions having superior dissolutionproperties.

Any suitable spray drying equipment may be used to co-dry the Reb Xsweetener composition and one or more co-agents. Those of ordinary skillin the art will appreciate that the equipment selection may be tailoredto obtain a product having particular physical characteristics. Forexample, foam spray drying may be used to produce low bulk densityproducts. Alternatively, a fluid bed may be attached to the exit of thespray dryer to produce a product having enhanced dissolution rates foruse in instant products. Examples of spray dryers include, but are notlimited to, co-current nozzle tower spray dryers, co-current rotaryatomizer spray dryers, counter-current nozzle tower spray dryers, andmixed-flow fountain nozzle spray dryers.

The resulting co-dried Reb X sweetener compositions may be furthertreated or separated using techniques well known to those of ordinaryskill in the art. For example, a desired particle size distribution canbe obtained by using screening techniques. Alternatively, the resultingco-dried Reb X sweetener compositions may undergo further processing,such as agglomeration.

Spray drying uses liquid feeds that can be atomized (e.g., slurries,solutions, and suspensions). Alternative methods of drying may beselected depending on the type of feed. For example, freeze drying andpan drying are capable of handling not only liquid feeds, as describedabove, but also wet cakes and pastes. Paddle dryers, such as high speedpaddle dryers, can accept slurries, suspensions, gels, and wet cakes.Vacuum drum drying methods, although primarily used with liquid feeds,have great flexibility in handling feeds having a wide range ofviscosities.

The resulting co-dried Reb X sweetener compositions have surprisingfunctionality for use in a variety of systems. Notably, the co-dried RebX sweetener compositions are believed to have superior taste properties.In addition, co-dried Reb X sweetener compositions may have increasedstability in low-moisture systems.

The present invention is further illustrated by the following example,which is not to be construed in any way as imposing limitations upon thescope thereof. On the contrary, it is to be clearly understood thatresort may be had to various other embodiments, modifications, andequivalents thereof which, after reading the description therein, maysuggest themselves to those skilled in the art without departing fromthe spirit of the present invention and/or the scope of the appendedclaims.

EXAMPLES Example 1 Purification of Reb X from Stevia rebaudiana BertoniPlant Leaves

Two kg of Stevia rebaudiana Bertoni plant leaves were dried at 45° C. toan 8.0% moisture content and ground to 10-20 mm particles. The contentof different glycosides in the leaves was as follows: Stevioside—2.55%,Reb A—7.78%, Reb B—0.01%, Reb C—1.04%, Reb D—0.21%, Reb F—0.14%, RebX—0.10% Dulcoside A—0.05%, and Steviolbioside—0.05%. The dried materialwas loaded into a continuous extractor and the extraction was carriedout with 40.0 L of water at a pH of 6.5 at 40° C. for 160 min. Thefiltrate was collected and subjected to chemical treatment. Calciumoxide in the amount of 400 g was added to the filtrate to adjust the pHwithin the range of 8.5-9.0, and the mixture was maintained for 15 minwith slow agitation. Then, the pH was adjusted to around 3.0 by adding600 g of FeCl₃ and the mixture was maintained for 15 min with slowagitation. A small amount of calcium oxide was further added to adjustthe pH to 8.5-9.0 and the mixture was maintained for 30 min with slowagitation. The precipitate was removed by filtration on aplate-and-frame filter press using cotton cloth as the filtrationmaterial. The slightly yellow filtrate was passed through the column,packed with cation-exchange resin Amberlite FCP22 (H⁺) and then, throughthe column with anion-exchange resin Amberlite FPA53 (OH⁻). The flowrate in both columns was maintained at SV=0.8 hour⁻¹. After completionboth columns were washed with RO water to recover the steviol glycosidesleft in the columns and the filtrates were combined. The portion ofcombined solution containing 120 g total steviol glycosides was passedthrough seven columns, wherein each column was packed with specificmacroporous polymeric adsorbent YWD-03 (Cangzhou Yuanwei, China). Thefirst column with the size of ⅓ of the others acted as a “catchercolumn”. The SV was around 1.0 hour⁻¹. After all extract was passedthrough the columns, the resin sequentially was washed with 1 volume ofwater, 2 volumes of 0.5% NaOH, 1 volume of water, 2 volumes of 0.5% HCl,and finally with water until the pH was 7.0. The “catcher column” waswashed separately. Desorption of the adsorbed steviol glycosides wascarried out with 52% ethanol at SV=1.0 hour⁻¹. Desorption of the first“catcher column” was carried out separately and the filtrate was notmixed with the main solution obtained from other columns. Desorption ofthe last column also was carried out separately. The quality of extractfrom different columns with specific macroporous adsorbent is shown inTable 1.

TABLE 1 Columns Total steviol glycosides, % 1 (catcher) 55.3 2 92.7 394.3 4 96.1 5 96.3 6 95.8 7 80.2

Eluates from second to sixth columns were combined and treatedseparately. The combined solution of steviol glycosides was mixed with0.3% of activated carbon from the total volume of solution. Thesuspension was maintained at 25° C. for 30 min with continuousagitation. Separation of carbon was carried out on a press-filtrationsystem. For additional decolorization the filtrate was passed throughthe columns packed with cation-exchange resin Amberlite FCP22 (H⁺)followed with anion-exchange resin Amberlite FPA53 A30B (OH⁻). The flowrate in both columns was around SV=0.5 hour⁻¹. The ethanol was distilledusing a vacuum evaporator. The solids content in the final solution wasaround 15%. The concentrate was passed through the columns packed withcation-exchange resin Amberlite FCP22 (H⁺) and anion-exchange resinAmberlite FPA53 (OH⁻) with SV=5 hour⁻¹. After all the solution waspassed through the columns, both resins were washed with RO water torecover the steviol glycosides left in the columns. The resultingrefined extract was transferred to the nano-filtration device,concentrated to around 52% of solids content and spray dried to providea highly purified mixture of steviol glycosides. The yield was 99.7 g.The mixture contained Stevioside—20.5%, Reb A—65.6%, Reb B—0.1%, RebC—8.4%, Reb D—0.5%, Reb F—1.1%, Reb X—0.1%, Dulcoside A—0.4%, andSteviolbioside—0.4%.

The combined eluate from the last column, contained about 5.3 g of totalsteviol glycosides including 2.3 g Reb D and around 1.9 g Reb X (35.8%Reb X/TSG ratio). It was deionized and decolorized as discussed aboveand then concentrated to a 33.5% content of total solids.

The concentrate was mixed with two volumes of anhydrous methanol andmaintained at 20-22° C. for 24 hours with intensive agitation.

The resulting precipitate was separated by filtration and washed withabout two volumes of absolute methanol. The yield of Reb X was 1.5 gwith around 80% purity.

For the further purification the precipitate was suspended in threevolumes of 60% methanol and treated at 55° C. for 30 min, then cooleddown to 20-22° C. and agitated for another 2 hours.

The resulting precipitate was separated by filtration and washed withabout two volumes of absolute methanol and subjected to similartreatment with a mixture of methanol and water.

The yield of Reb X was 1.2 g with 97.3% purity.

Example 2 Sensory Properties of Reb X

The sensory properties of Reb X were evaluated in acidified water (pH3.0 by phosphoric acid) at 500 mg/L concentration by 20 panelists. Theresults are summarized in Table 2.

TABLE 2 Evaluation of steviol glycosides at 500 ppm (pH 3.0) Number ofpanelists detected the attribute Stevioside Reb A Reb D Reb X SucroseTaste attribute (500 ppm) (500 ppm) (500 ppm) (500 ppm) (10,000 ppm)Bitter taste 20 20 3 0 0 Astringent taste 20 20 3 0 0 Licorice taste 2020 2 0 0 Sweet Aftertaste 20 20 5 0 0 Comments Quality of sweet BitterBitter Clean (9 of Clean (20 Clean (20 taste aftertaste aftertaste 20)of 20) of 20) (20 of 20) (20 of 20) Overall Satisfactory SatisfactorySatisfactory Satisfactory Satisfactory evaluation (0 of 20) (1 of 20)(11 of 20) (20 of 20) (20 of 20)

The above results clearly show that Reb X possesses superior tasteprofile to already known steviol glycosides.

Example 3 Structure Elucidation of Reb X

HRMS: HRMS (High Resolution Mass Spectrum) data was generated with aWaters Premier Quadrupole Time-of-Flight (Q-TOF) mass spectrometerequipped with an electrospray ionization source operated in thepositive-ion mode. Samples were diluted and eluted with a gradient of2:2:1 methanol:acetonitrile:water and introduced 50 μL via infusionusing the onboard syringe pump

NMR: The sample was dissolved in deuterated pyridine (C₅D₅N) and NMRspectra were acquired on Varian Unity Plus 600 MHz instruments usingstandard pulse sequences. The chemical shifts are given in δ (ppm), andcoupling constants are reported in Hz.

The complete ¹H and ¹³C NMR spectral assignments for the diterpeneglycoside rebaudioside X determined on the basis of 1D (′H and ¹³C) and2D (COSY, HMQC and HMBC) NMR as well as high resolution massspectroscopic data:

Discussion

The molecular formula was deduced as C₅₆H₉₀O₃₃ on the basis of itspositive high resolution (HR) mass spectrum (FIG. 6) which showed an[M+NH₄ ⁺] ion at mlz 1308.5703 together with an [M+Na+] adduct at mlz1313.5274. This composition was supported by ¹³C NMR spectral data (FIG.7). The ¹H NMR spectrum (FIG. 8) showed the presence of two methylsinglets at δ 1.32 and 1.38, two olefinic protons as singlets at δ 4.90and 5.69 of an exocyclic double bond, nine methylene and two methineprotons between δ 0.75-2.74 characteristic for the ent-kauranediterpenoids isolated earlier from the genus Stevia.

The basic skeleton of ent-kaurane diterpenoids was supported by COSY(FIG. 9): H-1/H-2; H-2/H-3; H-5/H-6; H-6/H-7; H-9/H-11; H-11/H-12correlations.

The basic skeleton of ent-kaurane diterpenoids was also supported byHMBC (FIG. 10): H-1/C-2, C-10; H-3/C-1, C-2, C-4, C-5, C-18, C-19;H-5/C-4, C-6, C-7, C-9, C-10, C-18, C-19, C-20; H-9/C-8, C-10, C-11,C-12, C-14, C-15; H-14/C-8, C-9, C-13, C-15, C-16 and H-17/C-13, C-15,C-16 correlations.

The ¹H NMR spectrum also showed the presence of anomeric protonsresonating at δ 5.31, 5.45, 5.46, 5.48, 5.81, and 6.39; suggesting sixsugar units in its structure. Enzymatic hydrolysis furnished an aglyconewhich was identified as steviol by comparison of co-TLC with standardcompound. Acid hydrolysis with 5% H₂SO₄ afforded glucose which wasidentified by direct comparison with authentic samples by TLC. The ¹Hand ¹³C NMR values for all protons and carbons were assigned on thebasis of COSY, HMQC and HMBC correlations (Table 3).

TABLE 3 ¹H and ¹³C NMR spectral data for Rebaudioside X in C₅D₅N^(a-c).Position ¹³C NMR ¹H NMR  1 40.3 0.75 t (13.2) 1.76 m  2 19.6 1.35 m 2.24m  3 38.4 1.01 m 2.30 d (13.3)  4 44.3 —  5 57.4 1.06 d (12.8)  6 23.52.23 m 2.41 q (13.2)  7 42.6 1.41 m 1.80 m  8 41.2 —  9 54.3 0.91 d(7.7) 10 39.7 — 11 20.2 1.65 m 1.75 m 12 38.5 1.86 m 2.73 m 13 87.6 — 1443.3 2.02 m 2.74 m 15 46.5 1.88 d (16.4) 2.03 m 16 153.3 — 17 104.9 4.90s 5.69 s 18 28.2 1.32 s 19 176.9 — 20 16.8 1.38 s  1′ 94.9 6.39 d (8.2) 2′ 76.9 4.51 t (8.5)  3′ 88.6 5.09 t (8.5)  4′ 70.1 4.18 m  5′ 78.44.13 m  6′ 61.8 4.20 m 4.31 m  1″ 96.2 5.46 d (7.1)  2″ 81.4 4.13 m  3″87.9 4.98 t (8.5)  4″ 70.4 4.07 t (9.6)  5″ 77.7 3.94 m  6″ 62.6 4.19 m4.32 m  1′″ 104.8 5.48 d (7.7)  2′″ 75.8 4.15 m  3′″ 78.6 4.13 m  4′″73.2 3.98 m  5′″ 77.6 3.74 ddd (2.8, 6.4, 9.9)  6′″ 64.0 4.27 m 4.51 m 1″″ 103.9 5.45 d (7.5)  2″″ 75.6 3.98 m  3″″ 77.8 4.50 t (7.8)  4″″71.3 4.14 m  5″″ 78.0 3.99 m  6″″ 62.1 4.20 m 4.32 m  1″″′ 104.2 5.81 d(7.2)  2″″′ 75.5 4.20 m  3″″′ 78.4 4.20 m  4″″′ 73.6 4.10 m  5″″′ 77.83.90 ddd (2.8, 6.4, 9.9)  6″″′ 64.0 4.32 m 4.64 d (10.3)  1″″″ 104.15.31 d (8.0)  2″″″ 75.5 3.95 m  3″″″ 78.0 4.37 t (9.1)  4″″″ 71.1 4.10 m 5″″″ 78.1 3.85 ddd (1.7, 6.1, 9.9)  6″″″ 62.1 4.10 m 4.32 m^(a)assignments made on the basis of COSY, HMQC and HMBC correlations;^(b)Chemical shift values are in δ (ppm); ^(c)Coupling constants are inHz.

Based on the results from NMR spectral data, it was concluded that thereare six glucosyl units. A close comparison of the ¹H and ¹³C NMRspectrum of Reb X with rebaudioside D suggested that Reb X was also asteviol glycoside which had three glucose residues that attached at theC-13 hydroxyl as a 2,3-branched glucotriosyl substituent and another2,3-branched glucotriosyl moiety in the form of an ester at C-19.

The key COSY and HMBC correlations suggested the placement of the sixthglucosyl moiety at the C-3 position of Sugar I. The large couplingconstants observed for the six anomeric protons of the glucose moietiesat δ 5.31 (d, J=8.0 Hz), 5.45 (d, J=7.5 Hz), 5.46 (d, J=7.1 Hz), 5.48(d, J=7.7 Hz), 5.81 (d, J=7.2 Hz), and 6.39 (d, J=8.2 Hz), suggestedtheir β-orientation as reported for steviol glycosides. Based on theresults of NMR and mass spectral studies and in comparison with thespectral values of rebaudioside A and rebaudioside D, Reb X was assignedas(β-[2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl)oxy]entkaur-16-en-19-oic acid-[2-O-β-D-glucopyranosyl-3-O-β-D-glycopyranosyl)ester.

Example 4 Taste Evaluation of Rebaudioside X

The taste properties of a sample of Reb X were studied againstRebaudioside A (Reb A) and Rebaudioside D (Reb D) samples. Reb A wasobtained from Cargill (lot#1040) and Reb-D was obtained from PureCircle(lot #11/3/08).

The samples were prepared at 500 ppm for sweetness evaluation by addingmoisture compensated mass into a 100 mL sample of carbon-treated waterand citric buffer solutions.

Citric buffer was prepared by mixing 1.171 g/L citric acid, 0.275 g/Lsodium citrate and 0.185 g/L sodium benzoate with carbon-treated water,with a final pH of 3.22. The mixtures were moderately stirred at roomtemperature. The Reb X sample was then evaluated against the two controlReb A and Reb D samples in water and citric buffer at room temperature(RT) and at 4° C. in an ice bath by one experienced panelist for anytasting quality determinations using the controlled, multi-sip andswallow taste method shown below:

1. Take first sip (˜1.8 mL) of control and swallow. Wait for 15-25 sec,then take second sip of control and wait for 15-25 sec.2. Take first sip of the experimental sample, wait for 15-25 sec, thentake second sip. Compare to second sip of control.3. Repeat steps #1 and #2 for the third and fourth sips of the controland experimental samples to confirm the initial finding.

Discussion:

The taste evaluation results of the Reb X samples against the controlReb A and Reb D samples at 500 ppm in citric buffer (CB) at 4° C. and RTare described in Table 4.

TABLE 4 Sample Taste Properties Reb A Delayed sweetness onset,significant lingering sweet (room aftertaste, licorice and bitteroff-notes. temperature) Reb X (room Clean sweet taste. Broad and fullsweetness profile. temperature) Moderate sweetness linger aftertastesimilar to aspartame. No bitter or licorice off-notes. Much bettertemporal profile than Reb A or Reb D. Smooth and broad sweetnesstemporal profile and quality similar to sucrose Reb D (room Delayedsweetness onset, less sweetness linger as temperature) compared to RebA, no bitter off-notes. Reb A Similar taste quality as Reb A at room butsignificantly (4° C.) more sweet Reb X Similar taste quality as Reb X atroom temperature (4° C.) Reb D Similar taste quality as Reb D at roombut significantly (4° C.) more sweet

The taste quality of Reb X at room temperature and 4° C. was similar.The taste quality of Reb X was much better than Reb A or Reb D. The RebX did not display a pure sugar-like tasting quality but rather containeda fatty-like or broader sweet temporal profile and less sweetness lingerthan that of Reb A. Similar to Reb D, Reb X did not have the astringencyor sweetness intense (depth) and bitterness notes compared to that ofthe Reb A in citrate buffer system.

Example 5 Solubility Studies of Reb X in Carbon-Treated Water andCitrate Buffer

The samples used to evaluate the taste properties in Example 2 were alsoused for solubility studies. At 500 ppm concentrations in citratebuffer, the initial solubility test revealed that Reb X has a similar,limited but significantly greater solubility than that of Reb D andsignificantly less solubility than Reb A.

Further solubility tests revealed the detailed data of concentration andtime to solubilize, as shown in Table 5:

TABLE 5 Estimated time to solubilize Reb X concentration in matrixsystem (minutes) 0.01% in carbon-treated water 15 0.02% incarbon-treated water 25 0.04% in carbon-treated water 33 0.05% incarbon-treated water 39 0.075% in carbon-treated water 55 0.10% incarbon-treated water 106  0.15% in carbon-treated water Insoluble after20 hours 0.01% in citrate buffer 25 0.02% in citrate buffer 25 0.04% incitrate buffer 35 0.05% in citrate buffer 42 0.075% in citrate buffer 550.10% in citrate buffer 106  0.15% in citrate buffer Insoluble after 20hours

Example 6 Isosweetness Determination of Reb X

The isosweetness levels of Reb X in a citric buffer system at roomtemperature and 4° C. were evaluated. A 600 ppm stock solution of Reb Xwas prepared by adding a mass of 0.15 g into a 250 mL sample of citricbuffer (CB) solution. The mixture was moderately stirred at warmertemperature (up to ca. 52° C.) on a heated stirrer for about 15-20minutes and then cooled. The citric buffer was prepared by adding 1.6 gcitric acid, 0.6 g potassium citrate and 0.253 g sodium benzoate in 1 Lof carbon-treated water. The pH of the mixture was 3.1. Seven dilutedReb X solutions at 12.5, 25, 50, 100, 200, 300, 400 and 500 ppm wereprepared by adding a 2.08, 4.17, 8.33, 16.67, 25.00, 33.33 and 41.67 mLReb X stock solution, respectively, into each 50 mL solution of CB. Thecontrols of 0.75%, 2%, 4%, 6%, 8%, 10% and 15% sucrose equivalence (SE)were also prepared by adding sugar (w/v) into the CB. The mixtures weremoderately stirred and then ready for the isosweetness determinationtest. The Reb X samples were then evaluated against the control sugarsamples in citric buffer at room temperature (RT) and 4° C. (in an icebath) by one experienced taster for an isosweetness determination usingthe controlled, multi-sip and swallow taste method. The results areshown in Table 6.

TABLE 6 Estimated % Estimated % Reb X Concentration SweetnessEquivalence Sweetness Equivalence (ppm) in Citric Buffer at RT in CitricBuffer at 4° C. 12.5 0.5  0.75 25 1.0-1.5 1.5 50 3.25-3.75 3.0-3.5 1005.0 5.5 200 8.0 8.0 300 12.0  11.0-12.0 400 14.0-14.5 14.0-14.5 50015.5-16.0 15.5-16.0 600 16.5-17.0 16.5-17.0

Discussion

The Reb X sample at 0.06% (w/v) was found to be very soluble and clear(colorless) in citric buffer at up to 52° C. for ca. 15-20 minutes. Nooff-flavors at any Reb X concentrations in CB at 4° C. were detected,except at least ca. 300 ppm which noticeably had a longer sweetnesslingering. At all concentrations a pleasant sweetness tasting qualitywith a slight delay of sweetness onset and no bitterness was detected.Despite the stronger mouthfeel or texture effect (syrupy, thicker) atca. 15% sucrose, it was difficult to determine the isosweetness levelsfor at least 400 ppm of Reb X due to its thinner mouthfeel but broaderand more impact sweetness temporal profile as well as its significantsweet lingering.

There was no significant sweetness intensity difference between RT and4° C. of the Reb X concentration range based on direct comparison withcontrol sucrose at similar temperatures. Two repeated tests comparing50, 100, 200, 300, 400 and 500 ppm Reb X concentrations at RT and 4° C.confirmed these initial results.

Example 7 Beverage Formulations Flavored Black Tea:

The taste properties of a flavored zero calorie black tea drinkcontaining Reb A in a concentration of 250 ppm was compared to acomparable flavored zero calorie black tea drink with Reb X in aconcentration of 250 ppm. The drink containing Reb X was determined tobe much cleaner in finish with less sweetness linger and a more roundedoverall sweetness profile.

Enhanced Water:

The taste properties of a zero calorie enhanced water drink containingReb A in a concentration of 200 ppm was compared to a comparable zerocalorie enhanced water drink containing Reb X in a concentration of 200ppm. The Reb X-containing drink was cleaner in finish and had reducedsweetness linger and a more rounded overall sweetness taste quality.

Orange-Flavored Sparking Beverage:

Reb X levels were evaluated in a zero calorie orange-flavored sparkingbeverage base to determine the effect of increasing sweetness. Samplesof the orange-flavored sparkling beverage with Reb X in amounts between400 and 750 ppm (in 50 ppm increments) were prepared. All samples tastedsignificantly better than comparable Reb A—containing formulationsresulting in cleaner profiles with increased sweetness intensity and nonegative aftertaste characteristic. Samples having 500 ppm and 550 ppmReb X were found to be the closest in sweetness level to a 11.5 Brixhigh fructose corn syrup sweetened orange flavored sparkling beverageformulation.

Example 8 Reb X Sweetness vs. Concentration

2.5%, 5.0%, 7.5%, and 10.0% sucrose solutions were prepared in neutral(7.0 pH) and acidified water (3.2 pH) as reference samples. Solutionscontaining Reb X (98% purity) were prepared to match the sweetness ofeach sucrose reference in neutral and acidified water. Samples weretasted and verified by a panel of trained tasters in water at roomtemperature.

TABLE 7 Sweetness Equivalent (SE) 2.5% 5.0% 7.5% 10.0% Reb XConcentration 48 132 254 422 (ppm) Sweetness Factor (SF) 521 380 295 237

Example 9 Sensory Comparison of Reb X and Reb A

To compare the sensory attributes between Reb X and Reb A, iso-sweetsamples having 8% sucrose equivalent sweetness were made with filteredwater as shown in Table 8. An 8% sugar solution in water at roomtemperature was used as a control.

TABLE 8 Reb A Formulation Reb X Formulation (weight percent of (weightpercent of Ingredient ingredient) ingredient) Water 99.95 99.95 Reb A(97% on a dry 0.0510 0 basis) Reb X (98% on a dry 0 0.0423 basis) Total100% 100%

Acidified solutions of 250 ppm citric acid (pH 3.2) containing the sameconcentration of Reb X and Reb A as indicated in Table 8 were alsoprepared. An 8% sugar solution in the acidified solution was used as thecontrol.

The samples prepared with filtered water were evaluated by 34semi-trained panel members at room temperature. The samples preparedwith acidified water were evaluated by 23 semi-trained panel members atroom temperature. Samples were given to the panel members sequentiallyand coded with triple digit numbers. The order of sample presentationwas randomized to avoid order of presentation bias. Water and unsaltedcrackers were provided in order to cleanse the palate. The panel memberswere asked to rate different attributes including sweetness onset, totalsweetness, rounded sweetness, bitterness, acidity, leafy note, licorice,astringency, mouthfeel, mouth coating, sweet lingering, and bitterlingering. Samples were rated on a scale of zero (0) to ten (10), withzero indicating immediate onset, no intensity, watery/low viscosity, orvery sharp peak, and ten indicating very delayed onset, high intensity,thick/high viscosity, or very round peak. One-way single factor ANOVAwas used to analyze sensory results, where α=0.05. The results are shownin FIGS. 11 and 12.

Discussion

Although Reb A and Reb X exhibited similar sweetness intensity, thefiltered water samples (FIG. 11) showed reduced perception ofbitterness, astringency and bitter lingering compared to Reb A. Inacidified water, the perception of higher sweetness of Reb X over Reb Ais significant (FIG. 12). Reb X also showed faster sweetness onset,reduced non-sweet taste (bitterness, sour, astringency) and bitternesslingering.

Example 10 Sensory Comparison of Reb X and Other Non-Caloric Sweeteners

Blends of Reb X and One Other Non-Caloric Sweetener

To study the interaction between Reb X and other natural ingredients,Reb X was blended with Reb B, Reb D, Reb A, NSF-02 (PureCirle),Mogroside V (Mog), and erythritol at various concentrations (Table 9) inacidified water and sensory evaluations were performed. The mainobjective of this study was to evaluate the improvement in the sweetnessprofile, including sweetness intensity in presence of otherco-ingredient/sweetener.

TABLE 9 RebX RebB RebD RebA Mog NSF02 Erythritol Sample (ppm) (ppm)(ppm) (ppm) (ppm) (ppm) (%) 1 300 2 300 100 3 300 50 4 300 50 50 5 300100 6 200 100 100 7 8 200 200 9 300 100 10 300 25 11 300 100 12 300 1%13 300 2%

The sweetened samples containing Reb X and Reb B were evaluated by 13semi-trained panel members at room temperature. The sweetened samplescontaining Reb X and NSF-02 were evaluated by 11 semi-trained panelmembers at room temperature. The sweetened samples containing Reb X andmogroside V were evaluated by 9 semi-trained panel members at roomtemperature. The sweetened samples containing Reb X and erythritol wereevaluated by 12 semi-trained panel members at room temperature. In allcases, samples were given to the panel members sequentially and codedwith triple digit numbers. The order of sample presentation wasrandomized to avoid order of presentation bias. Water and unsaltedcrackers were provided in order to cleanse the palate. The panel memberswere asked to rate different attributes including sweetness onset, totalsweetness, rounded sweetness, bitterness, acidity, leafy note, licorice,astringency, mouthfeel, mouth coating, sweet lingering, and bitterlingering. Samples were rated on a scale of zero (0) to ten (10), withzero indicating immediate onset, no intensity, watery/low viscosity, orvery sharp peak, and ten indicating very delayed onset, high intensity,thick/high viscosity, or very round peak. One-way single factor ANOVAwas used to analyze sensory results, where α=0.05. The results are shownin FIGS. 13-16.

Discussion

The Reb X/Reb B blends showed increased sweetness (i.e. synergy)compared to Reb X alone (FIG. 14). The Reb X/Reb B blend also showed amore rounded sweetness profile with improvement in sweetness intensity,onset and bitterness perception compared to Reb X alone.

The Reb X/NSF-02 blends had an overall rounded taste profile (FIG. 13).25 ppm NSF-02 shows a slight improvement in overall sweetness profileover Reb X alone, but had little impact on other attributes. The RebX/NSF-02 blend with 100 ppm NSF-02 had delayed sweetness onset and aslight increased sweetness intensity.

The Reb X/mogroside V blends had increased astringency, sourness andmouthcoating compared to the other blends evaluated (FIG. 15). Highermogroside V levels increased sweetness and sweetness lingering.

The Reb X/erythritol blends had an overall rounded taste profile (FIG.16). The blends had reduced acidity, reduced bitterness, reducedastringency and reduced bitterness lingering compared to Reb X alone. Atlevels above 1% (by weight), erythritol provides additional sweetnessand earlier sweetness onset.

Blends of Reb X and Two Other Non-Caloric Sweeteners

Three sets of the following formulations were prepared:

Formulation 1: 300 ppm Reb X

Formulation 2: 300 ppm Reb X and 100 ppm Reb A

Formulation 3: 300 ppm Reb X and 100 ppm Reb D

All samples were prepared in acidified water. The sweetened samples wereevaluated by 7 semi-trained panel members at room temperature. Sampleswere given to the panel members sequentially and coded with triple digitnumbers. The order of sample presentation was randomized to avoid orderof presentation bias. Water and unsalted crackers were provided in orderto cleanse the palate. The panel members were asked to rate differentattributes including sweetness onset, total sweetness, roundedsweetness, bitterness, acidity, leafy note, licorice, astringency,mouthfeel, mouth coating, sweet lingering, and bitter lingering. Sampleswere rated on a scale of zero (0) to ten (10), with zero indicatingimmediate onset, no intensity, watery/low viscosity, or very sharp peak,and ten indicating very delayed onset, high intensity, thick/highviscosity, or very round peak. One-way single factor ANOVA was used toanalyze sensory results, where α=0.05. The results are shown in FIG. 17.

Discussion

Both formulation 2 (Reb X and Reb A) and formulation 3 (Reb X and Reb D)showed increased total sweetness and overall sweetness profile(sweetness peak) compared to Reb X alone. In addition, both formulations2 and 3 showed decreased leafy note compared to Reb X alone. Formulation3 showed higher improvement in sweetness intensity, overall sweetnessprofile, bitter lingering and sweet lingering.

Blends of Reb X and Three Other Non-Caloric Sweeteners

Three sets of the following formulations were prepared:

Formulation 1: 300 ppm Reb X,

Formulation 2: 200 ppm Reb X, 100 ppm Reb A and 100 ppm Reb D

Formulation 3: 300 ppm Reb X, 50 ppm Reb B and 50 ppm Reb D.

All samples were prepared in acidified water. The sweetened samples wereevaluated by 11 semi-trained panel members at room temperature. Sampleswere given to the panel members sequentially and coded with triple digitnumbers. The order of sample presentation was randomized to avoid orderof presentation bias. Water and unsalted crackers were provided in orderto cleanse the palate. The panel members were asked to rate differentattributes including sweetness onset, total sweetness, roundedsweetness, bitterness, acidity, leafy note, licorice, astringency,mouthfeel, mouth coating, sweet lingering, and bitter lingering. Sampleswere rated on a scale of zero (0) to ten (10), with zero indicatingimmediate onset, no intensity, watery/low viscosity, or very sharp peak,and ten indicating very delayed onset, high intensity, thick/highviscosity, or very round peak. One-way single factor ANOVA was used toanalyze sensory results, where α=0.05. The results are shown in FIG. 18.

Discussion

Both formulation 2 (Reb X, Reb A and Reb D) and formulation 3 (Reb X,Reb B and Reb D) showed increased sweetness onset, overall sweetnessprofile (sweetness peak) and decreased lingering (bitter and sweetlingering) compared to Reb X alone. Formulation 2, which had lower Reb Xcontent compared to formulations 1 and 3, shoed a greater improvement inoverall sweetness profile and lingering.

1-61. (canceled)
 62. A method for purifying steviol glycosides, comprising the steps of: a. providing a solution comprising steviol glycosides; b. providing a plurality of consecutively connected columns packed with an adsorbent resin capable of adsorbing steviol glycosides to create a multi-column system; c. passing the solution of steviol glycosides through the multi-column system to obtain at least two columns, each column having a different ratio of adsorbed steviol glycosides; and d. eluting the adsorbed steviol glycosides from at least one column in the multi-column system, to obtain an eluted solution comprising steviol glycosides including Rebaudioside X.
 63. The method of claim 62, further comprising eluting high Rebaudioside X fractions from the multi-column system to obtain an eluted Rebaudioside X solution.
 64. The method of claim 62, further comprising the step of removing impurities by washing the multi-column system with a washing solution prior to eluting the adsorbed steviol glycosides.
 65. The method of claim 64, wherein the washing solution comprises an aqueous alcohol solution having a water to alcohol ratio ranging from about 99.9:0.1 to about 60:40.
 66. The method of claim 62, wherein the adsorbed steviol glycosides are eluted with an aqueous alcohol solution having a water to alcohol ratio ranging from about 60:40 to about 0.1:99.9.
 67. The method of claim 62, wherein 3 to 15 columns are consecutively connected in series.
 68. The method of claim 62, wherein a ratio of a volume of a first column of the plurality of columns to a volume of a second column of the plurality of columns is in the range of about 1:1 to about 1:10.
 69. The method of claim 62, wherein a ratio of a volume of a last column of the plurality of columns to a volume of a penultimate column of the plurality of columns is in the range of about 3:1 to about 1:10.
 70. The method of claim 62, wherein the solution of steviol glycosides is obtained by dissolving a composition comprising a steviol glycoside mixture in a solvent.
 71. The method of claim 62, further comprising the step of drying the eluted solution comprising steviol glycosides to obtain a purified steviol glycoside mixture.
 72. The method of claim 62, wherein the solution of steviol glycosides is prepared by a. providing a biomass of a Stevia rebaudiana plant; b. producing a crude extract by contacting the biomass with water; c. separating insoluble material from the crude extract to obtain a filtrate containing steviol glycosides; and d. treating the filtrate to remove high molecular weight compounds and insoluble particles to obtain the solution of steviol glycosides.
 73. The method of claim 62, further comprising the steps of: a. decolorizing and deionizing the eluted solution comprising steviol glycosides to obtain a decolorized, deionized steviol glycoside solution; and b. drying the decolorized, deionized steviol glycoside solution to obtain a purified steviol glycoside mixture.
 74. The method of claim 62, wherein steviol glycosides are selected from the group consisting of Rebaudioside A, Rebaudioside B, Rebaudioside C, Rebaudioside D, Rebaudioside E, Rebaudioside F, Rebaudioside X, Stevioside, Steviolbioside, Dulcoside A, Rubusoside, or other glycoside of steviol and combinations thereof.
 75. The method of claim 64, wherein the washing solution comprises at least one alkali.
 76. The method of claim 64, wherein the washing solution comprises at least one acid.
 77. The method of claim 64, wherein the washing solution comprises water. 